145 results on '"Morikis, D."'
Search Results
2. From atoms to systems: a cross-disciplinary approach to complement-mediated functions
- Author
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Mastellos, D, Morikis, D, Strey, C, Holland, C M., and Lambris, D J.
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- 2004
- Full Text
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3. Discovery of functionally selective C5aR2 ligands: novel modulators of C5a signalling
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Croker, D.E., Monk, P.N., Halai, R., Kaeslin, G., Schofield, Z., Wu, M.C., Clark, R.J., Blaskovich, M.A., Morikis, D., Floudas, C.A., Cooper, M.A., and Woodruff, T.M.
- Abstract
The complement cascade is comprised of a highly sophisticated network of innate immune proteins that are activated in response to invading pathogens or tissue injury. The complement activation peptide, C5a, binds two seven transmembrane receptors, namely the C5a receptor 1 (C5aR1) and C5a receptor 2 (C5aR2, or C5L2). C5aR2 is a non-G-protein-signalling receptor whose biological role remains controversial. Some of this controversy arises owing to the lack of selective ligands for C5aR2. In this study, a library of 61 peptides based on the C-terminus of C5a was assayed for the ability to selectively modulate C5aR2 function. Two ligands (P32 and P59) were identified as functionally selective C5aR2 ligands, exhibiting selective recruitment of β-arrestin 2 via C5aR2, partial inhibition of C5a-induced ERK1/2 activation and lipopolysaccharide-stimulated interleukin-6 release from human monocyte-derived macrophages. Importantly, neither ligand could induce ERK1/2 activation or inhibit C5a-induced ERK1/2 activation via C5aR1 directly. Finally, P32 inhibited C5a-mediated neutrophil mobilisation in wild-type, but not C5aR2(-/-) mice. These functionally selective ligands for C5aR2 are novel tools that can selectively modulate C5a activity in vitro and in vivo, and thus will be valuable tools to interrogate C5aR2 function.Immunology and Cell Biology advance online publication, 17 May 2016; doi:10.1038/icb.2016.43.
- Published
- 2016
4. Discovery of Small Molecules for Fluorescent Detection of Complement Activation Product C3d
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Gorham, RD, Nuñez, V, Lin, JH, Rooijakkers, SHM, Vullev, VI, and Morikis, D
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Protein Conformation ,1.1 Normal biological development and functioning ,Medicinal & Biomolecular Chemistry ,Organic Chemistry ,chemical and pharmacologic phenomena ,Pharmacology and Pharmaceutical Sciences ,Ligands ,Small Molecule Libraries ,Molecular Docking Simulation ,Structure-Activity Relationship ,Medicinal and Biomolecular Chemistry ,Underpinning research ,Complement C3d ,5.1 Pharmaceuticals ,Development of treatments and therapeutic interventions ,Complement Activation ,Fluorescent Dyes - Abstract
© 2015 American Chemical Society. Complement activation plays a major role in many acute and chronic inflammatory conditions. C3d, a terminal product of complement activation, remains covalently attached to cells and is an excellent biomarker of complement-mediated inflammation. We employed a virtual high-throughput screening protocol to identify molecules with predicted binding to complement C3d and with intrinsic fluorescence properties to enable detection. Pharmacophore models were developed based on known C3d-ligand interactions and information from computational analysis of structural and molecular dynamics data. Iterative pharmacophore-based virtual screening was performed to identify druglike molecules with physicochemical similarity to the natural C3d ligand CR2. Hits from the pharmacophore screens were docked to C3d and ranked based on predicted binding free energies. Top-ranked molecules were selected for experimental validation of binding affinity to C3d, using microscale thermophoresis, and for their suitability to become molecular imaging agents, using fluorescence spectroscopy. This work serves as a foundation for identifying additional fluorescent molecules with high-affinity for C3d that will subsequently be explored as noninvasive in vivo diagnostics of complement-mediated inflammation, for spatiotemporal monitoring of disease progression, and for targeting therapeutics to sites of inflammation.
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- 2015
5. Conformational analysis of compstatin analogues with molecular dynamics simulations in explicit water
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Tamamis, Phanourios, Skourtis, Spiros S., Morikis, D., Lambris, J. D., Archontis, Georgios Z., Tamamis, Phanourios [0000-0002-3342-2651], Skourtis, Spiros S. [0000-0002-5834-248X], and Archontis, Georgios Z. [0000-0002-7750-8641]
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Models, Molecular ,Computation theory ,Protein Conformation ,Mutant ,arginine ,Peptide ,Nuclear magnetic resonance ,Molecular dynamics ,Computational chemistry ,alpha helix ,Materials Chemistry ,Cluster Analysis ,Spectroscopy ,Physics ,chemistry.chemical_classification ,compstatin derivative ,education.field_of_study ,conformational transition ,Structural properties ,Intermolecular force ,article ,simulation ,Computer Graphics and Computer-Aided Design ,unclassified drug ,Conformations ,NMR spectra database ,priority journal ,Biochemistry ,Complement system ,Stereochemistry ,probability ,water ,Immunology ,Population ,Peptides, Cyclic ,peptide derivative ,Turn (biochemistry) ,Computer Simulation ,tryptophan ,Physical and Theoretical Chemistry ,education ,Molecular Biology ,Molecular dynamics simulations ,Water ,Compstatin analogues ,Conformational analysis ,chemistry ,Mutagenesis ,aspartic acid ,Helix ,Peptides ,cluster analysis - Abstract
The cyclic 13-residue peptide compstatin is a potential therapeutic agent against the unregulated activation of the complement system. A thorough knowledge of its structural and dynamical properties in solution may assist the design of improved complement inhibitors. NMR studies have suggested that the 5-8 segment of free compstatin folds into a critical for activity 5-8 β turn and the rest of the peptide is mainly disordered. Earlier computational studies of compstatin analogues with a polar-hydrogen/generalized-Born approximation reproduced the 5-8 turn, but also indicated the formation of β-hairpin or α-helical elements and the existence of interactions between certain charged or aromatic sidechains. However, these features are absent or partly present in the NMR spectra, due to extensive conformational averaging. In order to check the compstatin properties with a more rigorous model of the intra- and intermolecular interactions, we conduct here 98-ns all-atom/explicit-water simulations of three compstatin analogues with variable activity a native analogue, the more active mutant V4W/H9A and the inactive mutant Q5G. The 5-8 β-turn population is in good accord with NMR. For the systems studied here, the simulations suggest that the 5-8 turn population does not correlate strictly with activity, in agreement with earlier mutational studies. Furthermore, they show structural differences among the analogues outside the 5-8 region. The possible role of these differences in activity is discussed. The probability of β-hairpin or α-helix elements is much smaller with respect to the polar-hydrogen/GB simulations, and the persistent Trp4-Trp7 or Asp6-Arg11 sidechain interactions of the earlier GB studies are not reproduced. The present simulations extend the NMR data and improve our understanding of the properties of compstatin and related analogues. © 2007 Elsevier Inc. All rights reserved. 26 2 571 580 Cited By :10
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- 2007
6. Derivation of ligands for the complement C3a receptor from the C-terminus of C5a
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Halai, R., Bellows-Peterson, M.L., Branchett, W., Smadbeck, J., Kieslich, C.A., Croker, D.E., Cooper, M.A., Morikis, D., Woodruff, T.M., Floudas, C.A., and Monk, P.N.
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chemical and pharmacologic phenomena - Abstract
The complement cascade is a highly sophisticated network of proteins that are well regulated and directed in response to invading pathogens or tissue injury. Complement C3a and C5a are key mediators produced by this cascade, and their dysregulation has been linked to a plethora of inflammatory and autoimmune diseases. Consequently, this has stimulated interest in the development of ligands for the receptors for these complement peptides, C3a receptor, and C5a1 (C5aR/CD88). In this study we used computational methods to design novel C5a1 receptor ligands. However, functional screening in human monocyte-derived macrophages using the xCELLigence label-free platform demonstrated altered specificity of our ligands. No agonist/antagonist activity was observed at C5a1, but we instead saw that the ligands were able to partially agonize the closely related complement receptor C3a receptor. This was verified in the presence of C3a receptor antagonist SB 290157 and in a stable cell line expressing either C5a1 or C3a receptor alone. C3a agonism has been suggested to be a potential treatment of acute neutrophil-driven traumatic pathologies, and may have great potential as a therapeutic avenue in this arena.
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- 2014
7. Novel Compstatin Family Peptides Inhibit Complement Activation by Drusen-Like Deposits in Human Retinal Pigmented Epithelial Cell Cultures
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Gorham, R. D., Forest, D. L., Tamamis, Phanourios, López de Victoria, A., Kraszni, M., Kieslich, C. A., Banna, C. D., Bellows-Peterson, M. L., Larive, C. K., Floudas, C. A., Archontis, Georgios Z., Johnson, L. V., Morikis, D., Tamamis, Phanourios [0000-0002-3342-2651], and Archontis, Georgios Z. [0000-0002-7750-8641]
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Cell ,Peptide ,arginine ,protein binding ,Retinal Pigment Epithelium ,AMD ,C3b ,C3c ,chemistry.chemical_compound ,0302 clinical medicine ,lipophilicity ,retina macula age related degeneration ,protein data bank ,Complement Activation ,Compstatin family peptides ,Cells, Cultured ,apolipoprotein E ,chemistry.chemical_classification ,0303 health sciences ,C5b-9 ,article ,Sensory Systems ,peptide ,3. Good health ,unclassified drug ,pigment cell ,fetus ,medicine.anatomical_structure ,Biochemistry ,priority journal ,immunohistochemistry ,ELISA ,RPE ,reversed phase high performance liquid chromatography ,Complement inhibitors ,drug potency ,ApoE ,FB ,PDB ,in vitro study ,Complement system ,high performance liquid chromatography ,Retinal Drusen ,Drusen ,Biology ,Molecular dynamics ,Complement factor B ,Peptides, Cyclic ,Article ,complement component C3 ,03 medical and health sciences ,Cellular and Molecular Neuroscience ,peptide synthesis ,factor B ,medicine ,Humans ,controlled study ,human ,protein structure ,C3 ,the c-fragment of C3 ,age-related macular degeneration ,alternative pathway of complement activation ,030304 developmental biology ,nuclear magnetic resonance spectroscopy ,hydrophobicity ,complement activation ,cell culture ,Retinal pigment epithelium ,human cell ,Macular degeneration ,MD ,compstatin ,Retinal ,IC 50 ,the b-fragment of C3 ,medicine.disease ,biogenesis ,pigment epithelium ,eye diseases ,enzyme linked immunosorbent assay ,Ophthalmology ,complement system protein 3 ,the membrane attack complex consisting of complement proteins C5b, C6, C7, C8, and C9(n) ,chemistry ,concentration response ,RP-HPLC ,030221 ophthalmology & optometry ,drug solubility ,pathology ,enzyme-linked immunosorbent assay ,Retinal pigmented epithelium ,sense organs ,AP - Abstract
We have used a novel human retinal pigmented epithelial (RPE) cell-based model that mimics drusen biogenesis and the pathobiology of age-related macular degeneration to evaluate the efficacy of newly designed peptide inhibitors of the complement system. The peptides belong to the compstatin family and, compared to existing compstatin analogs, have been optimized to promote binding to their target, complement protein C3, and to enhance solubility by improving their polarity/hydrophobicity ratios. Based on analysis of molecular dynamics simulation data of peptide-C3 complexes, novel binding features were designed by introducing intermolecular salt bridge-forming arginines at the N-terminus and at position-1 of N-terminal dipeptide extensions. Our study demonstrates that the RPE cell assay has discriminatory capability for measuring the efficacy and potency of inhibitory peptides in a macular disease environment.© 2013 Elsevier Ltd. 116 96 108 Cited By :15
- Published
- 2013
8. De Novo Peptide Design with C3a Receptor Agonist and Antagonist Activities: Theoretical Predictions and Experimental Validation
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Bellows-Peterson, M.L., Fung, H.K., Floudas, C.A., Kieslich, C.A., Zhang, L., Morikis, D., Wareham, K.J., Monk, P.N., Hawksworth, O.A., and Woodruff, T.M.
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Targeting the complement component 3a receptor (C3aR) with selective agonists or antagonists is believed to be a viable therapeutic option for several diseases such as stroke, heart attack, reperfusion injuries, and rheumatoid arthritis. We designed a number of agonists, partial agonists, and antagonists of C3aR using our two-stage de novo protein design framework. Of the peptides tested using a degranulation assay in C3aR-transfected rat basophilic leukemia cells, two were prominent agonists (EC50 values of 25.3 and 66.2 nM) and two others were partial agonists (IC50 values of 15.4 and 26.1 nM). Further testing of these lead compounds in a calcium flux assay in U937 cells yielded similar results although with reduced potencies compared to transfected cells. The partial agonists also displayed full antagonist activity when tested in a C3aR inhibition assay. In addition, the electrostatic potential profile was shown to potentially discriminate between full agonists and partial agonists.
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- 2012
9. Molecular Dynamics in Drug Design: New Generations of Compstatin Analogs
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Tamamis, Phanourios, López de Victoria, A., Gorham, R. D., Bellows-Peterson, M. L., Pierou, P., Floudas, C. A., Morikis, D., Archontis, Georgios Z., Tamamis, Phanourios [0000-0002-3342-2651], and Archontis, Georgios Z. [0000-0002-7750-8641]
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crystal structure ,drug design ,Molecular Sequence Data ,Molecular modeling ,protein binding ,Molecular Dynamics Simulation ,Peptides, Cyclic ,hydroxymethylglutaryl coenzyme A reductase inhibitor ,Mice ,binding affinity ,physical chemistry ,Humans ,Animals ,Animalia ,rat ,Mechanism-based drug design ,human ,mouse ,hydrophobicity ,Complement system, C3 ,hydrogen bond ,nonhuman ,Rattus ,solubility ,article ,retina macula degeneration ,X ray crystallography ,molecular docking ,Complement C3 ,molecular dynamics ,amino acid sequence ,unclassified drug ,Rats ,priority journal ,protein stability ,protein protein interaction ,amino terminal sequence ,Compstatin ,protein secondary structure ,Structure-based drug design ,mutation - Abstract
We report the computational and rational design of new generations of potential peptide-based inhibitors of the complement protein C3 from the compstatin family. The binding efficacy of the peptides is tested by extensive molecular dynamics-based structural and physicochemical analysis, using 32 atomic detail trajectories in explicit water for 22 peptides bound to human, rat or mouse target protein C3, with a total of 257 ns. The criteria for the new design are: (i) optimization for C3 affinity and for the balance between hydrophobicity and polarity to improve solubility compared to known compstatin analogs and (ii) development of dual specificity, human-rat/mouse C3 inhibitors, which could be used in animal disease models. Three of the new analogs are analyzed in more detail as they possess strong and novel binding characteristics and are promising candidates for further optimization. This work paves the way for the development of an improved therapeutic for age-related macular degeneration, and other complement system-mediated diseases, compared to known compstatin variants. © 2012 John Wiley & Sons A/S. 79 5 703 718 Cited By :16
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- 2012
10. Design of a modified mouse protein with ligand binding properties of its human analog by molecular dynamics simulations: The case of C3 inhibition by compstatin
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Tamamis, Phanourios, Pierou, P., Mytidou, C., Floudas, C. A., Morikis, D., Archontis, Georgios Z., Tamamis, Phanourios [0000-0002-3342-2651], and Archontis, Georgios Z. [0000-0002-7750-8641]
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Innate immune response ,Models, Molecular ,Primates ,Complement system ,ligand binding ,Mice, Transgenic ,Molecular dynamics ,Molecular Dynamics Simulation ,Ligands ,Crystallography, X-Ray ,Peptides, Cyclic ,complement component C3 ,Mice ,protein conformation ,binding affinity ,Compstatin analogs ,Humans ,Animals ,Animalia ,Mus musculus ,human ,protein interaction ,Amino Acid Sequence ,protein structure ,Complement Activation ,mouse ,nonhuman ,Rattus ,article ,compstatin ,Complement C3 ,peptide ,unclassified drug ,priority journal ,Mammalia ,Peptides ,C3 inhibitors ,Sequence Alignment ,Protein Binding - Abstract
The peptide compstatin and its derivatives inhibit the complement-component protein C3 in primate mammals and are potential therapeutic agents against the unregulated activation of complement in humans, but are inactive against C3 from lower mammals. Recent molecular dynamics (MD) simulations showed that the most potent compstatin analog comprised entirely of natural amino acids (W4A9) had a smaller affinity for rat C3, due to reproducible changes in the rat protein structure with respect to the human protein, which eliminated or weakened specific protein-ligand interactions seen in the human C3:W4A9 complex. Here, we study by MD simulations three W4A9 complexes with the mouse C3 protein, and two "transgenic" mouse derivatives, containing a small number (6-9) of human C3 substitutions. The mouse complex experiences the conformational changes and affinity reduction of the rat complex. In the "transgenic" complexes, the conformation remains closer to that of the human complex, the protein-ligand interactions are improved, and the affinity for compstatin becomes "human-like." The present work creates new avenues for a compstatin-sensitive animal model. A similar strategy, involving the comparison of a series of complexes by MD simulations, could be used to design "transgenic" sequences in other systems. © 2011 Wiley-Liss, Inc. 79 11 3166 3179 Cited By :14
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- 2011
11. Electrostatic similarity determination using multiresolution analysis
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Hakkoymaz, H. Kieslich, C.A. Gorham Jr., R.D. Gunopulos, D. Morikis, D.
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Molecular similarity is an important tool in protein and drug design for analyzing the quantitative relationships between physicochemical properties of two molecules. We present a family of similarity measures which exploits the ability of wavelet transformation to analyze the spectral components of physicochemical properties and suggests a sensitive way for measuring similarities of biological molecules. In order to investigate how effective wavelet-based similarity measures were against conventional measures, we defined several patterns which involve scalar or topological changes in the distribution of electrostatic properties. The wavelet-based measures were more successful in discriminating these patterns in contrast to the current state-of-art similarity measures. We also present the validity of wavelet-based similarity measures through the hierarchical clustering of two protein datasets consisting of families of homologous domains and alanine scan mutants. This type of similarity analysis is useful for protein structure-function studies and protein design. Copyright © 2011 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.
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- 2011
12. Automated computational framework for the analysis of electrostatic similarities of proteins
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Kieslich, C.A. Morikis, D. Yang, J. Gunopulos, D.
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Charge plays an important role in protein-protein interactions. In the case of excessively charged proteins, their electrostatic potentials contribute to the processes of recognition and binding with other proteins or ligands. We present an automated computational framework for determining the contribution of each charged amino acid to the electrostatic properties of proteins, at atomic resolution level. This framework involves computational alanine scans, calculation of Poisson-Boltzmann electrostatic potentials, calculation of electrostatic similarity distances (ESDs), hierarchical clustering analysis of ESDs, calculation of solvation free energies of association, and visualization of the spatial distributions of electrostatic potentials. The framework is useful to classify families of mutants with similar electrostatic properties and to compare them with the parent proteins in the complex. The alanine scan mutants introduce perturbations in the local electrostatic properties of the proteins and aim in delineating the contribution of each mutated amino acid in the spatial distribution of electrostatic potential, and in biological function when electrostatics is a dominant contributing factor in protein-protein interactions. The framework can be used to design new proteins with tailored electrostatic properties, such as immune system regulators, inhibitors, and vaccines, and in guiding experimental studies. We present an example for the interaction of the immune system protein C3d (the d-fragment of complement protein C3) with its receptor CR2, and we discuss our data in view of a binding site controversy. © 2011 American Institute of Chemical Engineers.
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- 2011
13. Species specificity of the complement inhibitor compstatin investigated by all-atom molecular dynamics simulations
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Tamamis, Phanourios, Morikis, D., Floudas, C. A., Archontis, Georgios Z., Archontis, Georgios Z. [0000-0002-7750-8641], and Tamamis, Phanourios [0000-0002-3342-2651]
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Innate immune response ,Models, Molecular ,Primates ,Complement system ,X ray diffraction ,Protein Conformation ,Molecular Sequence Data ,Molecular Dynamics Simulation ,Peptides, Cyclic ,Species Specificity ,X-Ray Diffraction ,Compstatin analogs ,Humans ,Animals ,human ,protein interaction ,Amino Acid Sequence ,protein structure ,Complement Inactivator Proteins ,nonhuman ,Molecular dynamics simulations ,Rattus ,fungi ,article ,complement inhibitor ,compstatin ,Complement C3 ,simulation ,molecular dynamics ,unclassified drug ,Rats ,priority journal ,Mammalia ,mutation ,C3 inhibitors ,Sequence Alignment - Abstract
The development of compounds to regulate the activation of the complement system in non-primate species is of profound interest because it can provide models for human diseases. The peptide compstatin inhibits protein C3 in primate mammals and is a potential therapeutic agent against unregulated activation of complement in humans but is inactive against nonprimate species. Here, we elucidate this species specificity of compstatin by molecular dynamics simulations of complexes between the most potent natural compstatin analog and human or rat C3. The results are compared against an experimental conformation of the human complex, determined recently by X-ray diffraction at 2.4-å resolution. The human complex simulations provide information on the relative contributions to stability of specific C3 and compstatin residues. In the rat simulations, the protein undergoes reproducible conformational changes, which eliminate or weaken specific interactions and reduce the complex stability. The simulation insights can be used to design improved compstatin-based inhibitors for human C3 and active inhibitors against lower mammals. © 2010 Wiley-Liss, Inc. 78 12 2655 2667 Cited By :19
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- 2010
14. Engineering pre-SUMO4 as efficient substrate of SENP2
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Liu, Y., primary, Kieslich, C. A., additional, Morikis, D., additional, and Liao, J., additional
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- 2014
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15. The structural basis of compstatin activity examined by structure-function-based design of peptide analogs and NMR
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Morikis, D., Roy, M., Sahu, A., Troganis, A., Jennings, P. A., Tsokos, G. C., and Lambris, J. D.
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immunology ,biology ,2 parts ,complement inhibitor ,conformations ,activation ,c3 ,proteins - Abstract
We have previously identified compstatin, a 13-residue cyclic peptide, that inhibits complement activation by binding to C3 and preventing C3 cleavage to C3a and C3b. The structure of compstatin consists of a disulfide bridge and a type I beta-turn located at opposite sides to each other. The disulfide bridge is part of a hydrophobic cluster, and the beta-turn is part of a polar surface. We present the design of compstatin analogs in which we have introduced a series of perturbations in key structural elements of their parent peptide, compstatin. We have examined the consistency of the structures of the designed analogs compared with compstatin using NMR, and we have used the resulting structural information to make structure-complement inhibitory activity correlations. We propose the following. 1) Even in the absence of the disulfide bridge, a linear analog has a propensity for structure formation consistent with a turn of a 3(10)-helix or a beta-turn. 2) The type I beta-turn is a necessary but not a sufficient condition for activity. 3) Our substitutions outside the type I beta-turn of compstatin have altered the turn population but not the turn structure. 4) Flexibility of the beta-turn is essential for activity. 5) The type I beta-turn introduces reversibility and sufficiently separates the two sides of the peptide, whereas the disulfide bridge prevents the termini from drifting apart, thus aiding in the formation of the hydrophobic cluster. 6) The hydrophobic cluster at the linked termini is involved in binding to C3 and activity but alone is not sufficient for activity. 7) beta-Turn residues Gln(5) (Asn(5))-Asp(6)-Trp(7)(Phe(7)) -Gly(8) are specific for the turn formation, but only Gln(5)(Asn(5)) -Asp(6)-Trp(7)-Gly(8) residues are specific for activity. 8) Trp(7) is likely to be involved in direct interaction with C3, possibly through the formation of a hydrogen bond. Finally we propose a binding model for the C3-compstatin complex. Journal of Biological Chemistry
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- 2002
16. Insights into the mechanism of C5aR inhibition by PMX53 via implicit solvent molecular dynamics simulations and docking
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Tamamis, Phanourios, Kieslich, C. A., Nikiforovich, G. V., Woodruff, T. M., Morikis, D., Archontis, Georgios Z., Tamamis, Phanourios [0000-0002-3342-2651], and Archontis, Georgios Z. [0000-0002-7750-8641]
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C5a ,Complement system ,Molecular model ,Peptidomimetic ,1.1 Normal biological development and functioning ,Biophysics ,Nanotechnology ,Computational biology ,Molecular dynamics ,Implicit solvent ,C5a receptor ,Docking ,Underpinning research ,Anaphylatoxin ,C5aR ,Chemistry ,Intermolecular force ,Class A GPCR ,5.1 Pharmaceuticals ,Docking (molecular) ,Membrane protein ,Generic health relevance ,Development of treatments and therapeutic interventions ,Research Article - Abstract
Background: The complement protein C5a acts by primarily binding and activating the G-protein coupled C5a receptor C5aR (CD88), and is implicated in many inflammatory diseases. The cyclic hexapeptide PMX53 (sequence Ace-Phe-[Orn-Pro-dCha-Trp-Arg]) is a full C5aR antagonist of nanomolar potency, and is widely used to study C5aR function in disease. Results: We construct for the first time molecular models for the C5aR:PMX53 complex without the a priori use of experimental constraints, via a computational framework of molecular dynamics (MD) simulations, docking, conformational clustering and free energy filtering. The models agree with experimental data, and are used to propose important intermolecular interactions contributing to binding, and to develop a hypothesis for the mechanism of PMX53 antagonism. Conclusion: This work forms the basis for the design of improved C5aR antagonists, as well as for atomic-detail mechanistic studies of complement activation and function. Our computational framework can be widely used to develop GPCR-ligand structural models in membrane environments, peptidomimetics and other chemical compounds with potential clinical use. © 2014 Tamamis et al. licensee BioMed Central Ltd. 7 1 Cited By :7
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- 2014
17. New Compstatin Variants through Two De Novo Protein Design Frameworks
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Bellows, M.L., primary, Fung, H.K., additional, Taylor, M.S., additional, Floudas, C.A., additional, López de Victoria, A., additional, and Morikis, D., additional
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- 2010
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18. pH Dependence of Stability of the 10th Human Fibronectin Type III Domain: A Computational Study
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Mallik, B., primary, Zhang, L., additional, Koide, S., additional, and Morikis, D., additional
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- 2008
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19. Docking and dimerization domain (D/D) of the regulatory subunit of the Type II-alpha cAMP-dependent protein kinase A associated with a Peptide derived from an A-kinase anchoring protein (AKAP)
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Newlon, M.G., primary, Roy, M., additional, Morikis, D., additional, Hausken, Z.E., additional, Coghlan, V., additional, Scott, J.D., additional, and Jennings, P.A., additional
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- 2006
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20. Docking and dimerization domain (D/D) of the Type II-alpha regulatory subunity of protein kinase A (PKA) in complex with a peptide from an A-kinase anchoring protein
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Newlon, M.G., primary, Roy, M., additional, Morikis, D., additional, Hausken, Z.E., additional, Coghlan, V., additional, Scott, J.D., additional, and Jennings, P.A., additional
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- 2006
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21. Design of Peptide Analogues with Improved Activity Using a Novel de Novo Protein Design Approach
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Klepeis, J. L., primary, Floudas, C. A., additional, Morikis, D., additional, Tsokos, C. G., additional, and Lambris, J. D., additional
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- 2004
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22. Structural aspects and design of low-molecular-mass complement inhibitors
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Morikis, D., primary and Lambris, J. D., additional
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- 2002
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23. Rational design of a complement inhibitor targeting C3 using NMR and combinatorial approaches
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Morikis, D., primary, Soulika, A., additional, and Lambris, J.D., additional
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- 2002
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24. Solution structure of the docking and dimerization domain of protein kinase A II-alpha (RIIalpha D/D). Alternatively called the N-terminal dimerization domain of the regulatory subunit of protein kinase A.
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Morikis, D., primary, Roy, M., additional, Newlon, M.G., additional, Scott, J.D., additional, and Jennings, P.A., additional
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- 2002
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25. Predicting peptide structures using NMR data and deterministic global optimization
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Klepeis, J. L., primary, Floudas, C. A., additional, Morikis, D., additional, and Lambris, J. D., additional
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- 1999
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26. Hydration of the partially folded peptide RN-24 studied by multidimensional NMR
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Br�schweiler, R., primary, Morikis, D., additional, and Wright, P.E., additional
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- 1995
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27. Low pH myoglobin photoproducts
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Sage, J.T., primary, Morikis, D., additional, Li, P., additional, and Champion, P.M., additional
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- 1992
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28. Conformational interconversion in protein crystals
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Zhu, L., primary, Sage, J.T., additional, Rigos, A.A., additional, Morikis, D., additional, and Champion, P.M., additional
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- 1992
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29. Resonance Raman scattering as a probe of electron-nuclear coupling: applications to heme proteins
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Morikis, D., primary, Li, P., additional, Bangcharoenpaurpong, O., additional, Sage, J. T., additional, and Champion, P. M., additional
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- 1991
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30. Resonance Raman studies of iron spin and axial coordination in distal pocket mutants of ferric myoglobin.
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Morikis, D, primary, Champion, P M, additional, Springer, B A, additional, Egebey, K D, additional, and Sligar, S G, additional
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- 1990
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31. Design and NMR Characterization of Active Analogues of Compstatin Containing Non-Natural Amino Acids
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Mallik, B., Katragadda, M., Spruce, L. A., Carafides, C., Tsokos, C. G., Morikis, D., and Lambris, J. D.
- Abstract
We present new findings in our drug discovery effort to develop an anticomplement therapeutic. We have designed several active analogues of compstatin by altering its amino acid composition at positions 4 and 9. The most effective analogues have tryptophan or fused-ring non-natural amino acids at position 4 and alanine or an unbranched single-methyl amino acid at position 9. Twenty-one of these analogues have 2−99-fold higher activities compared to the parent peptide compstatin. The analogue Ac-V4(2Nal)/H9A-NH
2 has the highest inhibitory activity with IC50 500 nM. NMR data, through NOE and chemical shift analysis, suggest the presence of interconverting conformers spanning the extended and helical regions of the Ramachandran plot, and they detect a predominant averaged conformer with coil structure and at least one flexible β-turn, of type I. The fused-ring non-natural amino acids at position 4 contribute to the formation of the hydrophobic cluster of compstatin, which has been previously proposed, together with the β-turn and a disulfide bridge, to be essential for binding to the target of compstatin, complement component C3. We propose that additional mechanisms may contribute to the structural stability of the analogues and to binding to C3, involving intra- and intermolecular electrostatic interactions of the π-electron system of side chain aromatic rings. The presence of π−π interactions for Trp4−Trp7 was confirmed with a molecular dynamics simulation for the most active analogue with natural amino acids, Ac-V4W/H9A-NH2 . Alanine or aminobutyric acid at position 9 contribute to the weak propensity for helical structure of the residue segment 4−10 of the analogues, which may also play a role in increased activity.- Published
- 2005
32. Improvement of the anti-C3 activity of compstatin using rational and combinatorial approaches
- Author
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Morikis, D., Soulika, A.M., Mallik, B., Klepeis, J.L., Floudas, C.A., and Lambris, J.D.
- Abstract
Compstatin is a 13-residue cyclic peptide that has the ability to inhibit the cleavage of C3 to C3a and C3b. The effects of targeting C3 cleavage are threefold, and result in hindrance of: (i) the generation of the pro-inflammatory peptide C3a, (ii) the generation of opsonin C3b (or its fragment C3d), and (iii) further complement activation of the common pathway (beyond C3) with the end result of the generation of the membrane attack complex. We will report on our progress on: (i) rational design of more active compstatin analogues based on the three-dimensional structure of compstatin, (ii) experimental combinatorial design based on the generation of a phage-displayed peptide library partially randomized with the implementation of structure-induced restraints, and (iii) theoretical combinatorial design based on a novel computational optimization method, structure-induced restraints and flexible structural templates. All three approaches have resulted in analogues with improved activities. Currently, the lead analogue has the sequence acetyl-I[CVYQDWGAHRC]T-NH2 (where the brackets denote cyclization), and is 16-fold more active than the parent peptide. We will also report on our progress towards understanding the dynamic character of compstatin using molecular dynamics simulations. The identification of an ensemble of interconverting conformers of compstatin with variable populations is a first step towards the incorporation of dynamic elements in the design of new analogues using dynamics–activity relationships in addition to structure–activity relationships.
- Published
- 2004
- Full Text
- View/download PDF
33. Peptide redesign for inhibition of the complement system: Targeting age-related macular degeneration
- Author
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Rr, Mohan, Ap, Cabrera, Re, Harrison, Ronald Gorham, Lv, Johnson, Ghosh K, and Morikis D
34. Binding kinetics, structure-activity relationship, and biotransformation of the complement inhibitor Compstatin
- Author
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Sahu, A., Soulika, A. M., Morikis, D., Spruce, L., Moore, W. T., and John Lambris
35. Design of peptide analogues with improved activity using a novel de novo protein design approach
- Author
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Klepeis, J. L., Floudas, C. A., Morikis, D., Tsokos, C. G., and John Lambris
36. Systems Biology Modeling of the Complement System Under Immune Susceptible Pathogens.
- Author
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Zewde NT, Hsu RV, Morikis D, and Palermo G
- Abstract
The complement system is assembled from a network of proteins that function to bring about the first line of defense of the body against invading pathogens. However, complement deficiencies or invasive pathogens can hijack complement to subsequently increase susceptibility of the body to infections. Moreover, invasive pathogens are increasingly becoming resistant to the currently available therapies. Hence, it is important to gain insights into the highly dynamic interaction between complement and invading microbes in the frontlines of immunity. Here, we developed a mathematical model of the complement system composed of 670 ordinary differential equations with 328 kinetic parameters, which describes all three complement pathways (alternative, classical, and lectin) and includes description of mannose-binding lectin, collectins, ficolins, factor H-related proteins, immunoglobulin M, and pentraxins. Additionally, we incorporate two pathogens: (type 1) complement susceptible pathogen and (type 2) Neisseria meningitidis located in either nasopharynx or bloodstream. In both cases, we generate time profiles of the pathogen surface occupied by complement components and the membrane attack complex (MAC). Our model shows both pathogen types in bloodstream are saturated by complement proteins, whereas MACs occupy <<1.0% of the pathogen surface. Conversely, the MAC production in nasopharynx occupies about 1.5-10% of the total N. meningitidis surface, thus making nasal MAC levels at least about eight orders of magnitude higher. Altogether, we predict complement-imbalance, favoring overactivation, is associated with nasopharynx homeostasis. Conversely, orientating toward complement-balance may cause disruption to the nasopharynx homeostasis. Thus, for sporadic meningococcal disease, our model predicts rising nasal levels of complement regulators as early infection biomarkers., Competing Interests: Conflict of Interest: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.
- Published
- 2021
- Full Text
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37. Role of Electrostatic Hotspots in the Selectivity of Complement Control Proteins Toward Human and Bovine Complement Inhibition.
- Author
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Narkhede YB, Gautam AK, Hsu RV, Rodriguez W, Zewde NT, Harrison RES, Arantes PR, Gaieb Z, Gorham RD Jr, Kieslich C, Morikis D, Sahu A, and Palermo G
- Abstract
Poxviruses are dangerous pathogens, which can cause fatal infection in unvaccinated individuals. The causative agent of smallpox in humans, variola virus , is closely related to the bovine vaccinia virus , yet the molecular basis of their selectivity is currently incompletely understood. Here, we examine the role of the electrostatics in the selectivity of the smallpox protein SPICE and vaccinia protein VCP toward the human and bovine complement protein C3b, a key component of the complement immune response. Electrostatic calculations, in-silico alanine-scan and electrostatic hotspot analysis, as introduced by Kieslich and Morikis ( PLoS Comput. Biol . 2012), are used to assess the electrostatic complementarity and to identify sites resistant to local perturbation where the electrostatic potential is likely to be evolutionary conserved. The calculations suggest that the bovine C3b is electrostatically prone to selectively bind its VCP ligand. On the other hand, the human isoform of C3b exhibits a lower electrostatic complementarity toward its SPICE ligand. Yet, the human C3b displays a highly preserved electrostatic core, which suggests that this isoform could be less selective in binding different ligands like SPICE and the human Factor H. This is supported by experimental cofactor activity assays revealing that the human C3b is prone to bind both SPICE and Factor H, which exhibit diverse electrostatic properties. Additional investigations considering mutants of SPICE and VCP that revert their selectivity reveal an "electrostatic switch" into the central modules of the ligands, supporting the critical role of the electrostatics in the selectivity. Taken together, these evidences provide insights into the selectivity mechanism of the complement regulator proteins encoded by the variola and vaccinia viruses to circumvent the complement immunity and exert their pathogenic action. These fundamental aspects are valuable for the development of novel vaccines and therapeutic strategies., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2021 Narkhede, Gautam, Hsu, Rodriguez, Zewde, Harrison, Arantes, Gaieb, Gorham, Kieslich, Morikis, Sahu and Palermo.)
- Published
- 2021
- Full Text
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38. SPARC coordinates extracellular matrix remodeling and efficient recruitment to and migration of antigen-specific T cells in the brain following infection.
- Author
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McGovern KE, Nance JP, David CN, Harrison RES, Noor S, Worth D, Landrith TA, Obenaus A, Carson MJ, Morikis D, and Wilson EH
- Subjects
- Animals, Antigens, Protozoan immunology, Biomarkers, Brain blood supply, Brain immunology, Brain metabolism, Brain parasitology, Cell Movement immunology, Chemokine CCL21 metabolism, Disease Models, Animal, Epitopes, T-Lymphocyte immunology, Gene Expression Regulation, Host-Parasite Interactions immunology, Mice, Mice, Knockout, Neurons metabolism, Osteonectin genetics, Protein Binding, Receptors, CCR7, Extracellular Matrix metabolism, Osteonectin metabolism, T-Lymphocytes immunology, T-Lymphocytes metabolism, Toxoplasma physiology, Toxoplasmosis, Cerebral etiology, Toxoplasmosis, Cerebral metabolism
- Abstract
Central nervous system (CNS) injury and infection can result in profound tissue remodeling in the brain, the mechanism and purpose of which is poorly understood. Infection with the protozoan parasite Toxoplasma gondii causes chronic infection and inflammation in the brain parenchyma. Control of parasite replication requires the continuous presence of IFNγ-producing T cells to keep T. gondii in its slowly replicating cyst form. During infection, a network of extracellular matrix fibers, revealed using multiphoton microscopy, forms in the brain. The origin and composition of these structures are unknown but the fibers have been observed to act as a substrate for migrating T cells. In this study, we show a critical regulator of extracellular matrix (ECM) remodeling, Secreted Protein, Acidic, Rich in Cysteine (SPARC), is upregulated in the brain during the early phases of infection in the frontal cortex. In the absence of SPARC, a reduced and disordered fibrous network, increased parasite burden, and reduced antigen-specific T cell entry into the brain points to a role for SPARC in T cell recruitment to and migration within the brain. We also report SPARC can directly bind to CCR7 ligands CCL19 and CCL21 but not CXCL10, and enhance migration toward a chemokine gradient. Measurement of T cell behavior points to tissue remodeling being important for access of immune cells to the brain and facilitating cellular locomotion. Together, these data identify SPARC as an important regulatory component of immune cell trafficking and access to the inflamed CNS.
- Published
- 2021
- Full Text
- View/download PDF
39. Factor H-Inspired Design of Peptide Biomarkers of the Complement C3d Protein.
- Author
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Harrison RES, Zewde NT, Narkhede YB, Hsu RV, Morikis D, Vullev VI, and Palermo G
- Abstract
C3d is a hallmark protein of the complement system, whose presence is critical to measure the progression of several immune diseases. Here, we propose to directly target C3d through small peptides mimicking the binding of its natural ligand, the complement regulator Factor H (FH). Through iterative computational analysis and binding affinity experiments, we establish a rationale for the structure-based design of FH-inspired peptides, leading to low-micromolar affinity for C3d and stable binding over microsecond-length simulations. Our FH-inspired peptides call now for further optimization toward high-affinity binding and suggest that small peptides are promising as novel C3d biomarkers and therapeutic tools., Competing Interests: The authors declare no competing financial interest., (Copyright © 2020 American Chemical Society.)
- Published
- 2020
- Full Text
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40. Crosslinked flagella as a stabilized vaccine adjuvant scaffold.
- Author
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Gries CM, Mohan RR, Morikis D, and Lo DD
- Subjects
- Bacterial Proteins chemistry, Bacterial Proteins immunology, Cross-Linking Reagents chemistry, Disulfides, Flagella chemistry, Flagellin chemistry, Flagellin immunology, Models, Molecular, Protein Conformation, Protein Multimerization, Salmonella typhimurium chemistry, Salmonella typhimurium genetics, Bacterial Proteins metabolism, Bacterial Vaccines immunology, Flagella metabolism, Flagellin metabolism, Salmonella typhimurium metabolism
- Abstract
Background: Engineered vaccine proteins incorporating both antigen and adjuvant components are constructed with the aim of combining functions to induce effective protective immunity. Bacterial flagellin is a strong candidate for an engineered vaccine scaffold as it is known to provide adjuvant activity through its TLR5 and inflammasome activation. Moreover, polymerized flagellin filaments can elicit a more robust immunoglobulin response than monomeric flagellin, and the multimeric antigen form can also promote T cell-independent antibody responses. Here, we aim to produce and test a covalently stabilized polymerized flagellar filament, providing additional immune efficacy through stabilization of its polymeric filament structure, as well as stabilization for long-term storage., Results: Computational modeling of monomer packing in flagellin filaments helped identify amino acids with proximity to neighboring flagella protofilaments. Paired cysteine substitutions were made at amino acids predicted to form inter-monomer disulfide cross-links, and these substitutions were capable of forming flagella when transfected into a flagellin-negative strain of Salmonella enterica subspecies Typhimurium. Interestingly, each paired substitution stabilized different helical conformational polymorphisms; the stabilized filaments lost the ability to transition between conformations, reducing bacterial motility. More importantly, the paired substitutions enabled extensive disulfide cross links and intra-filament multimer formation, and in one of the three variants, permitted filament stability in high acidic and temperature conditions where wild-type filaments would normally rapidly depolymerize. In addition, with regard to potential adjuvant activity, all crosslinked flagella filaments were able to induce wild-type levels of epithelial NF-κB in a cell reporter system. Finally, bacterial virulence was unimpaired in epithelial adherence and invasion, and the cysteine substitutions also appeared to increase bacterial resistance to oxidizing and reducing conditions., Conclusions: We identified amino acid pairs, with cysteine substitutions, were able to form intermolecular disulfide bonds that stabilized the resulting flagellar filaments in detergent, hydrochloric acid, and high temperatures while retaining its immunostimulatory function. Flagellar filaments with disulfide-stabilized protofilaments introduce new possibilities for the application of flagella as a vaccine adjuvant. Specifically, increased stability and heat tolerance permits long-term storage in a range of temperature environments, as well as delivery under a range of clinical conditions.
- Published
- 2019
- Full Text
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41. Characterization, Dynamics, and Mechanism of CXCR4 Antagonists on a Constitutively Active Mutant.
- Author
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Rosenberg EM Jr, Harrison RES, Tsou LK, Drucker N, Humphries B, Rajasekaran D, Luker KE, Wu CH, Song JS, Wang CJ, Murphy JW, Cheng YC, Shia KS, Luker GD, Morikis D, and Lolis EJ
- Subjects
- Benzylamines, Chemokine CXCL12 pharmacology, Cyclams, HEK293 Cells, HIV Infections metabolism, HIV Infections pathology, HIV Infections virology, HIV-1 drug effects, Heterocyclic Compounds pharmacology, Humans, Hydrophobic and Hydrophilic Interactions, Ligands, Mutagenesis, Site-Directed, Protein Conformation, alpha-Helical, Protein Structure, Tertiary, Receptors, CXCR4 genetics, Receptors, CXCR4 metabolism, Signal Transduction drug effects, Small Molecule Libraries chemistry, Small Molecule Libraries pharmacology, beta-Arrestin 2 metabolism, Molecular Dynamics Simulation, Receptors, CXCR4 antagonists & inhibitors, Small Molecule Libraries metabolism
- Abstract
The G protein-coupled receptor (GPCR) CXCR4 is a co-receptor for HIV and is involved in cancers and autoimmune diseases. We characterized five purine or quinazoline core polyamine pharmacophores used for targeting CXCR4 dysregulation in diseases. All were neutral antagonists for wild-type CXCR4 and two were biased antagonists with effects on β-arrestin-2 only at high concentrations. These compounds displayed various activities for a constitutively active mutant (CAM). We use the IT1t-CXCR4 crystal structure and molecular dynamics (MD) simulations to develop two hypotheses for the activation of the N119
3.35 A CAM. The N1193.35 A mutation facilitates increased coupling of TM helices III and VI. IT1t deactivates the CAM by disrupting the coupling between TM helices III and VI, mediated primarily by residue F872.53 . Mutants of F872.53 in N1193.35 A CXCR4 precluded constitutive signaling and prevented inverse agonism. This work characterizes CXCR4 ligands and provides a mechanism for N1193.35 A constitutive activation., (Copyright © 2019 Elsevier Ltd. All rights reserved.)- Published
- 2019
- Full Text
- View/download PDF
42. Molecular Mechanisms of Macular Degeneration Associated with the Complement Factor H Y402H Mutation.
- Author
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Harrison RES and Morikis D
- Subjects
- Binding Sites, Complement Factor H genetics, Complement Factor H metabolism, Heparin chemistry, Humans, Protein Binding, Complement Factor H chemistry, Macular Degeneration genetics, Molecular Docking Simulation, Molecular Dynamics Simulation, Mutation, Missense
- Abstract
A single nucleotide polymorphism, tyrosine at position 402 to histidine (Y402H), within the gene encoding complement factor H (FH) predisposes individuals to acquiring age-related macular degeneration (AMD) after aging. This polymorphism occurs in short consensus repeat (SCR) 7 of FH and results in decreased binding affinity of SCR6-8 for heparin. As FH is responsible for regulating the complement system, decreased affinity for heparin results in decreased regulation on surfaces of self. To understand the involvement of the Y402H polymorphism in AMD, we leverage methods from bioinformatics and computational biophysics to quantify structural and dynamical differences between SCR7 isoforms that contribute to decreased pattern recognition in SCR7
H402 . Our data from molecular and Brownian dynamics simulations suggest a revised mechanism for decreased heparin binding. In this model, transient contacts not observed in structures for SCR7 are predicted to occur in molecular dynamics simulations between coevolved residues Y402 and I412, stabilizing SCR7Y402 in a conformation that promotes association with heparin. H402 in the risk isoform is less likely to form a contact with I412 and samples a larger conformational space than Y402. We observe energy minima for sidechains of Y402 and R404 from SCR7Y402 that are predicted to associate with heparin at a rate constant faster than energy minima for sidechains of H402 and R404 from SCR7H402 . As both carbohydrate density and degree of sulfation decrease with age in Bruch's membrane of the macula, the decreased heparin recognition of SCR7H402 may contribute to the pathogenesis of AMD., (Copyright © 2018 Biophysical Society. Published by Elsevier Inc. All rights reserved.)- Published
- 2019
- Full Text
- View/download PDF
43. M13 bacteriophage spheroids as scaffolds for directed synthesis of spiky gold nanostructures.
- Author
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Ngo-Duc TT, Plank JM, Chen G, Harrison RES, Morikis D, Liu H, and Haberer ED
- Abstract
The spherical form (s-form) of a genetically-modified gold-binding M13 bacteriophage was investigated as a scaffold for gold synthesis. Repeated mixing of the phage with chloroform caused a 15-fold contraction from a nearly one micron long filament to an approximately 60 nm diameter spheroid. The geometry of the viral template and the helicity of its major coat protein were monitored throughout the transformation process using electron microscopy and circular dichroism spectroscopy, respectively. The transformed virus, which retained both its gold-binding and mineralization properties, was used to assemble gold colloid clusters and synthesize gold nanostructures. Spheroid-templated gold synthesis products differed in morphology from filament-templated ones. Spike-like structures protruded from the spherical template while isotropic particles developed on the filamentous template. Using inductively coupled plasma-mass spectroscopy (ICP-MS), gold ion adsorption was found to be comparatively high for the gold-binding M13 spheroid, and likely contributed to the dissimilar gold morphology. Template contraction was believed to modify the density, as well as the avidity of gold-binding peptides on the scaffold surface. The use of the s-form of the M13 bacteriophage significantly expands the templating capabilities of this viral platform and introduces the potential for further morphological control of a variety of inorganic material systems.
- Published
- 2018
- Full Text
- View/download PDF
44. Dissecting Distinct Roles of NEDDylation E1 Ligase Heterodimer APPBP1 and UBA3 Reveals Potential Evolution Process for Activation of Ubiquitin-related Pathways.
- Author
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Malik-Chaudhry HK, Gaieb Z, Saavedra A, Reyes M, Kung R, Le F, Morikis D, and Liao J
- Subjects
- Fluorescence Resonance Energy Transfer, Humans, Molecular Dynamics Simulation, NEDD8 Protein genetics, NEDD8 Protein metabolism, Protein Binding, Protein Domains, Protein Subunits chemistry, Protein Subunits genetics, Protein Subunits metabolism, Static Electricity, Ubiquitin-Activating Enzymes genetics, Ubiquitin-Activating Enzymes metabolism, Evolution, Molecular, NEDD8 Protein chemistry, Ubiquitin-Activating Enzymes chemistry
- Abstract
Despite the similar enzyme cascade in the Ubiquitin and Ubiquitin-like peptide(Ubl) conjugation, the involvement of single or heterodimer E1 activating enzyme has been a mystery. Here, by using a quantitative Förster Resonance Energy Transfer (FRET) technology, aided with Analysis of Electrostatic Similarities Of Proteins (AESOP) computational framework, we elucidate in detail the functional properties of each subunit of the E1 heterodimer activating-enzyme for NEDD8, UBA3 and APPBP1. In contrast to SUMO activation, which requires both subunits of its E1 heterodimer AOS1-Uba2 for its activation, NEDD8 activation requires only one of two E1 subunits, UBA3. The other subunit, APPBP1, only contributes by accelerating the activation reaction rate. This discovery implies that APPBP1 functions mainly as a scaffold protein to enhance molecular interactions and facilitate catalytic reaction. These findings for the first time reveal critical new mechanisms and a potential evolutionary pathway for Ubl activations. Furthermore, this quantitative FRET approach can be used for other general biochemical pathway analysis in a dynamic mode.
- Published
- 2018
- Full Text
- View/download PDF
45. Virtual Screening of Chemical Compounds for Discovery of Complement C3 Ligands.
- Author
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Mohan RR, Wilson M, Gorham RD Jr, Harrison RES, Morikis VA, Kieslich CA, Orr AA, Coley AV, Tamamis P, and Morikis D
- Abstract
The complement system is our first line of defense against foreign pathogens, but when it is not properly regulated, complement is implicated in the pathology of several autoimmune and inflammatory disorders. Compstatin is a peptidic complement inhibitor that acts by blocking the cleavage of complement protein C3 to the proinflammatory fragment C3a and opsonin fragment C3b. In this study, we aim to identify druglike small-molecule complement inhibitors with physicochemical, geometric, and binding properties similar to those of compstatin. We employed two approaches using various high-throughput virtual screening methods, which incorporate molecular dynamics (MD) simulations, pharmacophore model design, energy calculations, and molecular docking and scoring. We have generated a library of 274 chemical compounds with computationally predicted binding affinities for the compstatin binding site of C3. We have tested subsets of these chemical compounds experimentally for complement inhibitory activity, using hemolytic assays, and for binding affinity, using microscale thermophoresis. As a result, although none of the compounds showed inhibitory activity, compound 29 was identified to exhibit weak competitive binding against a potent compstatin analogue, therefore validating our computational approaches. Additional docking and MD simulation studies suggest that compound 29 interacts with C3 residues, which have been shown to be important in binding of compstatin to the C3c fragment of C3. Compound 29 is amenable to physicochemical optimization to acquire inhibitory properties. Additionally, it is possible that some of the untested compounds will demonstrate binding and inhibition in future experimental studies., Competing Interests: The authors declare the following competing financial interest(s): D.M., P.T., R.D.G., R.R.M., and R.E.S.H. are coinventors in a patent and/or patent application(s) for compstatin family peptides.
- Published
- 2018
- Full Text
- View/download PDF
46. A computational model for the evaluation of complement system regulation under homeostasis, disease, and drug intervention.
- Author
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Zewde N and Morikis D
- Subjects
- Atypical Hemolytic Uremic Syndrome diagnosis, Atypical Hemolytic Uremic Syndrome immunology, Biomarkers analysis, Biomarkers metabolism, Complement Inactivating Agents pharmacology, Complement Inactivating Agents therapeutic use, Complement Pathway, Alternative drug effects, Complement Pathway, Classical drug effects, Complement System Proteins genetics, Complement System Proteins immunology, Computational Biology, Glomerulonephritis, Membranoproliferative diagnosis, Glomerulonephritis, Membranoproliferative immunology, Homeostasis drug effects, Homeostasis physiology, Humans, Immunity, Innate drug effects, Immunity, Innate physiology, Macular Degeneration diagnosis, Macular Degeneration immunology, Complement Pathway, Alternative physiology, Complement Pathway, Classical physiology, Complement System Proteins metabolism, Models, Biological
- Abstract
The complement system is an intricate defense network that rapidly removes invading pathogens. Although many complement regulators are present to protect host cells under homeostasis, the impairment of Factor H (FH) regulatory mechanism has been associated with several autoimmune and inflammatory diseases. To understand the dynamics involved in the pivotal balance between activation and regulation, we have developed a comprehensive computational model of the alternative and classical pathways of the complement system. The model is composed of 290 ordinary differential equations with 142 kinetic parameters that describe the state of complement system under homeostasis and disorder through FH impairment. We have evaluated the state of the system by generating concentration-time profiles for the biomarkers C3, C3a-desArg, C5, C5a-desArg, Factor B (FB), Ba, Bb, and fC5b-9 that are influenced by complement dysregulation. We show that FH-mediated disorder induces substantial levels of complement activation compared to homeostasis, by generating reduced levels of C3 and FB, and to a lesser extent C5, and elevated levels of C3a-desArg, Ba, Bb, C5a-desArg, and fC5b-9. These trends are consistent with clinically observed biomarkers associated with complement-mediated diseases. Furthermore, we introduced therapy states by modeling known inhibitors of the complement system, a compstatin variant (C3 inhibitor) and eculizumab (C5 inhibitor). Compstatin demonstrates strong restorative effects for early-stage biomarkers, such as C3a-desArg, FB, Ba, and Bb, and milder restorative effects for late-stage biomarkers, such as C5a-desArg and fC5b-9, whereas eculizumab has strong restorative effects on late-stage biomarkers, and negligible effects on early-stage biomarkers. These results highlight the need for patient-tailored therapies that target early complement activation at the C3 level, or late-stage propagation of the terminal cascade at the C5 level, depending on the specific FH-mediated disease and the manifestations of a patient's genetic profile in complement regulatory function., Competing Interests: The authors have received a research grant from Achillion Pharmaceuticals, a company that specializes on complement-based drug discovery, for a disease-specific computational model. DM has received lecture/travel honorarium from Achillion Pharmaceuticals. DM is a co-inventor in a patent on compstatin peptides (Title: Compstatin Analogs; US Patent 9512180), and in provisional and patent applications on compstatin peptides. This does not alter our adherence to PLOS ONE policies on sharing data and materials.
- Published
- 2018
- Full Text
- View/download PDF
47. Conformational heterogeneity in CCR7 undergoes transitions to specific states upon ligand binding.
- Author
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Gaieb Z and Morikis D
- Subjects
- Binding Sites, Humans, Hydrogen Bonding, Ligands, Molecular Dynamics Simulation, Protein Binding, Protein Conformation, alpha-Helical, Receptors, CCR7 chemistry
- Abstract
Ligand-binding to G protein-coupled receptors (GPCRs) acts as the local driving force that initiates signal transduction through the receptor and mediates its conformational transitions and interactions with various intracellular effectors. In a recent study, We have shown that the binding of ligands CCL19 and CCL21 to CCR7 induces biased triggering of side chain-based molecular switches, which coordinate concerted transmembrane helical domain motions and transitioning of the receptor to distinct conformational states (Gaieb, Z., D.D. Lo, and D. Morikis. 2016. Molecular Mechanism of Biased Ligand Conformational Changes in CC Chemokine Receptor 7. Journal of Chemical Information and Modeling. 56: 1808-1822, DOI: 10.1021/acs.jcim.6b00367). To complement our previous study, we compare the results of the free (apo) CCR7 microsecond molecular dynamics simulations to those of the ligand-bound CCR7, and show that the apo receptor is found in conformational heterogeneity that only exhibits random fluctuations and lacks the coordinated helical motions seen in ligand-bound receptors. We conclude that ligand binding is responsible for coordinating the stochastic conformational nature of CCR7 into specific conformational states, initiated and propagated by specific physicochemical events., (Copyright © 2017 Elsevier Inc. All rights reserved.)
- Published
- 2017
- Full Text
- View/download PDF
48. AESOP: A Python Library for Investigating Electrostatics in Protein Interactions.
- Author
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Harrison RES, Mohan RR, Gorham RD Jr, Kieslich CA, and Morikis D
- Subjects
- Alanine chemistry, Alanine metabolism, Algorithms, Internet, Mutation, Proteins genetics, Proteins metabolism, Thermodynamics, Proteins chemistry, Software, Static Electricity
- Abstract
Electric fields often play a role in guiding the association of protein complexes. Such interactions can be further engineered to accelerate complex association, resulting in protein systems with increased productivity. This is especially true for enzymes where reaction rates are typically diffusion limited. To facilitate quantitative comparisons of electrostatics in protein families and to describe electrostatic contributions of individual amino acids, we previously developed a computational framework called AESOP. We now implement this computational tool in Python with increased usability and the capability of performing calculations in parallel. AESOP utilizes PDB2PQR and Adaptive Poisson-Boltzmann Solver to generate grid-based electrostatic potential files for protein structures provided by the end user. There are methods within AESOP for quantitatively comparing sets of grid-based electrostatic potentials in terms of similarity or generating ensembles of electrostatic potential files for a library of mutants to quantify the effects of perturbations in protein structure and protein-protein association., (Copyright © 2017 Biophysical Society. Published by Elsevier Inc. All rights reserved.)
- Published
- 2017
- Full Text
- View/download PDF
49. Ionic tethering contributes to the conformational stability and function of complement C3b.
- Author
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López-Perrote A, Harrison RE, Subías M, Alcorlo M, Rodríguez de Córdoba S, Morikis D, and Llorca O
- Subjects
- Animals, Complement C3b genetics, Genetic Predisposition to Disease, Humans, Microscopy, Electron, Polymorphism, Single Nucleotide, Protein Conformation, Protein Domains physiology, Protein Stability, Thermodynamics, Complement C3b chemistry, Complement C3b metabolism, Macular Degeneration genetics, Models, Molecular
- Abstract
C3b, the central component of the alternative pathway (AP) of the complement system, coexists as a mixture of conformations in solution. These conformational changes can affect interactions with other proteins and complement regulators. Here we combine a computational model for electrostatic interactions within C3b with molecular imaging to study the conformation of C3b. The computational analysis shows that the TED domain in C3b is tethered ionically to the macroglobulin (MG) ring. Monovalent counterion concentration affects the magnitude of electrostatic forces anchoring the TED domain to the rest of the C3b molecule in a thermodynamic model. This is confirmed by observing NaCl concentration dependent conformational changes using single molecule electron microscopy (EM). We show that the displacement of the TED domain is compatible with C3b binding to Factor B (FB), suggesting that the regulation of the C3bBb convertase could be affected by conditions that promote movement in the TED domain. Our molecular model also predicts mutations that could alter the positioning of the TED domain, including the common R102G polymorphism, a risk variant for developing age-related macular degeneration. The common C3b isoform, C3bS, and the risk isoform, C3bF, show distinct energetic barriers to displacement in the TED that are related to a network of electrostatic interactions at the interface of the TED and MG-ring domains of C3b. These computational predictions agree with experimental evidence that shows differences in conformation observed in C3b isoforms purified from homozygous donors. Altogether, we reveal an ionic, reversible attachment of the TED domain to the MG ring that may influence complement regulation in some mutations and polymorphisms of C3b., (Copyright © 2016 Elsevier Ltd. All rights reserved.)
- Published
- 2017
- Full Text
- View/download PDF
50. Detection of Side Chain Rearrangements Mediating the Motions of Transmembrane Helices in Molecular Dynamics Simulations of G Protein-Coupled Receptors.
- Author
-
Gaieb Z and Morikis D
- Abstract
Structure and dynamics are essential elements of protein function. Protein structure is constantly fluctuating and undergoing conformational changes, which are captured by molecular dynamics (MD) simulations. We introduce a computational framework that provides a compact representation of the dynamic conformational space of biomolecular simulations. This method presents a systematic approach designed to reduce the large MD simulation spatiotemporal datasets into a manageable set in order to guide our understanding of how protein mechanics emerge from side chain organization and dynamic reorganization. We focus on the detection of side chain interactions that undergo rearrangements mediating global domain motions and vice versa. Side chain rearrangements are extracted from side chain interactions that undergo well-defined abrupt and persistent changes in distance time series using Gaussian mixture models, whereas global domain motions are detected using dynamic cross-correlation. Both side chain rearrangements and global domain motions represent the dynamic components of the protein MD simulation, and are both mapped into a network where they are connected based on their degree of coupling. This method allows for the study of allosteric communication in proteins by mapping out the protein dynamics into an intramolecular network to reduce the large simulation data into a manageable set of communities composed of coupled side chain rearrangements and global domain motions. This computational framework is suitable for the study of tightly packed proteins, such as G protein-coupled receptors, and we present an application on a seven microseconds MD trajectory of CC chemokine receptor 7 (CCR7) bound to its ligand CCL21.
- Published
- 2017
- Full Text
- View/download PDF
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