Search

For the last several years, searching of new xanthone derivatives (XDs) with potential pharmacological activities has remained one of the main areas of interest of our group. The optimization of biological activity and drug-like properties of hits and leads is crucial at early stage of the drug discovery pipeline. Lipophilicity is one of the most important drug-like properties having a great impact in both pharmacokinetics and pharmacodynamics processes. In this work, we describe the lipophilicity of a small library of bioactive XDs, previously synthesized by our group, using different methods: computational, vortex-assisted liquid–liquid microextraction coupled with high-performance liquid chromatography (VALLME-HPLC), reversed-phase high-performance thin layer chromatography (RP-HPTLC), reversed-phase high-performance liquid chromatography (RP-HPLC), and biomembrane model by the partition between micelles and aqueous phase. The different results obtained by the used methods were compared and discussed. The methodologies and data gathered in this study will expand the investigation of lipophilicity of XDs, an important class of compounds in medicinal chemistry. © 2017 Elsevier This work was partially supported through national funds provided by FCT/MCTES − Foundation for Science and Technology from the Minister of Science, Technology and Higher Education (PIDDAC) and European Regional Development Fund (ERDF) through the COMPETE − Programa Operacional Factores de Competitividade (POFC) programme , under the Strategic Funding UID/Multi/04423/2013, the project PTDC/MAR-BIO/4694/2014 (reference POCI-01-0145-FEDER-016790; Project 3599–Promover a Produção Científica e Desenvolvimento Tecnológico e a Constituição de Redes Temáticas (3599-PPCDT)) in the framework of the programme PT2020 . José Soares thanks the financial support of National Funds from FCT (Fundação para a Ciência e a Tecnologia), FEDER under Program PT2020 (project 007265 −UID/QUI/50006/2013), and through the FCT PhD Programmes and by Programa Operacional Potencial Humano (POCH), specifically by the BiotechHealth Programme (Doctoral Programme on Cellular and Molecular Biotechnology Applied to Health Sciences), reference PD/00016/2012. José Soares thanks FCT and POPH (Programa Operacional Potencial Humano) for his PhD grant (SFRH/BD/98105/2013). Appendix A

We report a first set of peptidomimetic ligands mimicking the adhesive interface identified by recent crystallographic structures of E- and N-cadherin. Compounds 2 and 3 inhibit adhesion of epithelial ovarian cancer (EOC) cells with improved efficacy compared to the ADH-1 peptide, a N-cadherin antagonist that is in early clinical trials in EOC patients.

Fibrillar aggregates of the protein α-synuclein (αS) are one of the hallmarks of Parkinson’s disease. Here, we show that measuring the fluorescence polarization (FP) of labels at several sites on αS allows one to monitor changes in the local dynamics of the protein after binding to micelles or vesicles, and during fibril formation. Most significantly, these site-specific FP measurements provide insight into structural remodeling of αS fibrils by small molecules and have the potential for use in moderate-throughput screens to identify small molecules that could be used to treat Parkinson’s disease. © 2016 American Chemical Society.

The ASCT2 transport system catalyses a sodium-dependent antiport of glutamine and other neutral amino acids which is involved in amino acid metabolism. A library of 1,2,3-dithiazoles was designed, synthesized and evaluated as inhibitors of the glutamine/amino acid ASCT2 transporter in the model system of proteoliposomes reconstituted with the rat liver transporter. Fifteen of the tested compounds at concentration of 20μM or below, inhibited more than 50% the glutamine/glutamine antiport catalysed by the reconstituted transporter. These good inhibitors bear a phenyl ring with electron withdrawing substituents. The inhibition was reversed by 1,4-dithioerythritol indicating that the effect was likely owed to the formation of mixed sulfides with the protein's Cys residue(s). A dose-response analysis of the most active compounds gave IC50 values in the range of 3-30μM. Kinetic inhibition studies indicated a non-competitive inhibition, presumably because of a potential covalent interaction of the dithiazoles with cysteine thiol groups that are not located at the substrate binding site. Indeed, computational studies using a homology structural model of ASCT2 transporter, suggested as possible binding targets, Cys-207 or Cys-210, that belong to the CXXC motif of the protein. © 2012 Elsevier Inc. 265 1 93 102 Cited By :23

The genetic missense A30P mutation of the wild-type α-synuclein protein results in the replacement of the 30th amino acid residue from alanine (Ala) to proline (Pro) and was initially found in the members of a German family who developed Parkinson's disease. Even though the structures of these proteins have been measured before, detailed understanding about the structures and their relationships with free energy landscapes is lacking, which is of interest to provide insights into the pathogenic mechanism of Parkinson's disease. We report the secondary and tertiary structures and conformational free energy landscapes of the wild-type and A30P mutant-type α-synuclein proteins in an aqueous solution environment via extensive parallel tempering molecular dynamics simulations along with thermodynamic calculations. In addition, we present the residual secondary structure component transition stabilities at the atomic level with dynamics in terms of free energy change calculations using a new strategy that we reported most recently. Our studies yield new interesting results; for instance, we find that the A30P mutation has local as well as long-range effects on the structural properties of the wild-type α-synuclein protein. The helical content at Ala18-Gly31 is less prominent in comparison to the wild-type α-synuclein protein. The β-sheet structure abundance decreases in the N-terminal region upon A30P mutation of the wild-type α-synuclein, whereas the NAC and C-terminal regions possess larger tendencies for β-sheet structure formation. Long-range intramolecular protein interactions are less abundant upon A30P mutation, especially between the NAC and C-terminal regions, which is linked to the less compact and less stable structures of the A30P mutant-type rather than the wild-type α-synuclein protein. Results including the usage of our new strategy for secondary structure transition stabilities show that the A30P mutant-type α-synuclein tendency toward aggregation is higher than the wild-type α-synuclein but we also find that the C-terminal and NAC regions of the A30P mutant-type α-synuclein are reactive toward fibrillzation and aggregation based on atomic level studies with dynamics in an aqueous solution environment. Therefore, we propose that small molecules or drugs blocking the specific residues, which we report herein, located in the NAC- and C-terminal regions of the A30P mutant-type α-synuclein protein might help to reduce the toxicity of the A30P mutant-type α-synuclein protein. © 2013 American Chemical Society.

Single-domain antibodies (sdAbs) derived from human V H are considered to be less soluble and prone to aggregate which makes it difficult to determine the crystal structures. In this study, we isolated and characterized two anti-human epidermal growth factor receptor-2 (HER2) sdAbs, Gr3 and Gr6, from a synthetic human V H phage display library. Size exclusion chromatography and surface plasmon resonance analyses demonstrated that Gr3 is a monomer, but that Gr6 is a strict dimer. To understand this different molecular behavior, we solved the crystal structure of Gr6 to 1.6 Å resolution. The crystal structure revealed that the homodimer assembly of Gr6 closely mimics the V H-V L heterodimer of immunoglobulin variable domains and the dimerization interface is dominated by hydrophobic interactions. © 2012 Baral et al.

The ability of a peptide to target a specific protein in vitro has a potential for recognizing the cell membrane structure, protein-protein and receptor-ligand interaction. On the basis of molecular interactions, cell specific peptides were selected via phage library approach and their functions on the cells were determined by XTT based viability assay. We aimed to find phage displayed peptides from 12-mer peptide library that interact with K562-dox cell membrane in association with cellular functions and to study the effects of peptides selected from artificial peptide library on K562-dox cell viability. Peptides recognizing K562-dox cells were identified according to peptide sequences and amino acid properties. We selected 29 different phages from biopannings with K562-dox cells. Three clones were identified (KPB7,KPB10, KPP8) and their negative effects on cell viability were determined by XTT assay. According to our cell viability assay results, selected phages were effected negatively to the viability of the K562-dox cells. Depending on the present results, peptides were obtained that could be potential candidates for molecular recognition and cell targeting approaches. © 2010 Academic Journals Inc.