19 results on '"Ranade, Ranae M."'
Search Results
2. Structure-guided design of novel Trypanosoma brucei Methionyl-tRNA synthetase inhibitors
- Author
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Huang, Wenlin, Zhang, Zhongsheng, Barros-Álvarez, Ximena, Koh, Cho Yeow, Ranade, Ranae M., Gillespie, J. Robert, Creason, Sharon A., Shibata, Sayaka, Verlinde, Christophe L.M.J., Hol, Wim G.J., Buckner, Frederick S., and Fan, Erkang
- Published
- 2016
- Full Text
- View/download PDF
3. Identification of Potent Inhibitors of the Trypanosoma brucei Methionyl-tRNA Synthetase via High-Throughput Orthogonal Screening
- Author
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Pedró-Rosa, Laura, Buckner, Frederick S., Ranade, Ranae M., Eberhart, Christina, Madoux, Franck, Gillespie, J. Robert, Koh, Cho Yeow, Brown, Steven, Lohse, Jacqueline, Verlinde, Christophe L.M., Fan, Erkang, Bannister, Thomas, Scampavia, Louis, Hol, Wim G.J., Spicer, Timothy, and Hodder, Peter
- Published
- 2015
- Full Text
- View/download PDF
4. Methionyl-tRNA synthetase inhibitor has potent in vivo activity in a novel Giardia lamblia luciferase murine infection model.
- Author
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Michaels, Samantha A, Shih, Han-Wei, Zhang, Bailin, Navaluna, Edelmar D, Zhang, Zhongsheng, Ranade, Ranae M, Gillespie, J Robert, Merritt, Ethan A, Fan, Erkang, Buckner, Frederick S, Paredez, Alexander R, and Ojo, Kayode K
- Subjects
GIARDIASIS ,HIGH throughput screening (Drug development) ,ENZYMES ,GIARDIA lamblia ,OXIDOREDUCTASES ,MICE ,ANIMALS - Abstract
Background: Methionyl-tRNA synthetase (MetRS) inhibitors are under investigation for the treatment of intestinal infections caused by Giardia lamblia.Objectives: To properly analyse the therapeutic potential of the MetRS inhibitor 1717, experimental tools including a robust cell-based assay and a murine model of infection were developed based on novel strains of G. lamblia that employ luciferase reporter systems to quantify viable parasites.Methods: Systematic screening of Giardia-specific promoters and luciferase variants led to the development of a strain expressing the click beetle green luciferase. Further modifying this strain to express NanoLuc created a dual reporter strain capable of quantifying parasites in both the trophozoite and cyst stages. These strains were used to develop a high-throughput cell assay and a mouse infection model. A library of MetRS inhibitors was screened in the cell assay and Compound-1717 was tested for efficacy in the mouse infection model.Results: Cell viability in in vitro compound screens was quantified via bioluminescence readouts while infection loads in mice were monitored with non-invasive whole-animal imaging and faecal analysis. Compound-1717 was effective in clearing mice of Giardia infection in 3 days at varying doses, which was supported by data from enzymatic and phenotypic cell assays.Conclusions: The new in vitro and in vivo assays based on luciferase expression by engineered G. lamblia strains are useful for the discovery and development of new therapeutics for giardiasis. MetRS inhibitors, as validated by Compound-1717, have promising anti-giardiasis properties that merit further study as alternative therapeutics. [ABSTRACT FROM AUTHOR]- Published
- 2020
- Full Text
- View/download PDF
5. Triazolopyrimidines and Imidazopyridines: Structure–Activity Relationships and in Vivo Efficacy for Trypanosomiasis.
- Author
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Nagendar, Pendem, Gillespie, J. Robert, Herbst, Zackary M., Ranade, Ranae M., Molasky, Nora M. R., Faghih, Omeed, Turner, Rachael M., Gelb, Michael H., and Buckner, Frederick S.
- Published
- 2019
- Full Text
- View/download PDF
6. The crystal structure of the drug target <italic>Mycobacterium tuberculosis</italic> methionyl‐tRNA synthetase in complex with a catalytic intermediate.
- Author
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Barros-Álvarez, Ximena, Turley, Stewart, Ranade, Ranae M., Gillespie, J. Robert, Duster, Nicole A., Verlinde, Christophe L. M. J., Fan, Erkang, Buckner, Frederick S., and Hol, Wim G. J.
- Subjects
MYCOBACTERIUM tuberculosis ,CRYSTAL structure ,LIGASES - Abstract
Mycobacterium tuberculosis is a pathogenic bacterial infectious agent that is responsible for approximately 1.5 million human deaths annually. Current treatment requires the long‐term administration of multiple medicines with substantial side effects. Lack of compliance, together with other factors, has resulted in a worrisome increase in resistance. New treatment options are therefore urgently needed. Here, the crystal structure of methionyl‐tRNA synthetase (MetRS), an enzyme critical for protein biosynthesis and therefore a drug target, in complex with its catalytic intermediate methionyl adenylate is reported. Phenylalanine 292 of theM. tuberculosis enzyme is in an `out' conformation and barely contacts the adenine ring, in contrast to other MetRS structures where ring stacking occurs between the adenine and a protein side‐chain ring in the `in' conformation. A comparison with human cytosolic MetRS reveals substantial differences in the active site as well as regarding the position of the connective peptide subdomain 1 (CP1) near the active site, which bodes well for arriving at selective inhibitors. Comparison with the human mitochondrial enzyme at the amino‐acid sequence level suggests that arriving at inhibitors with higher affinity for the mycobacterial enzyme than for the mitochondrial enzyme might be achievable. [ABSTRACT FROM AUTHOR]- Published
- 2018
- Full Text
- View/download PDF
7. Urea Derivatives of 2-Aryl-benzothiazol-5-amines: A New Class of Potential Drugs for Human African Trypanosomiasis.
- Author
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Patrick, Donald A., Gillespie, J. Robert, McQueen, Joshua, Hulverson, Matthew A., Ranade, Ranae M., Creason, Sharon A., Herbst, Zackary M., Gelb, Michael H., Buckner, Frederick S., and Tidwell, Richard R.
- Published
- 2017
- Full Text
- View/download PDF
8. Brucella melitensis Methionyl-tRNA-Synthetase (MetRS), a Potential Drug Target for Brucellosis.
- Author
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Ojo, Kayode K., Ranade, Ranae M., Zhang, Zhongsheng, Dranow, David M., Myers, Janette B., Choi, Ryan, Nakazawa Hewitt, Steve, Edwards, Thomas E., Davies, Douglas R., Lorimer, Donald, Boyle, Stephen M., Barrett, Lynn K., Buckner, Frederick S., Fan, Erkang, and Van Voorhis, Wesley C.
- Subjects
- *
BRUCELLA melitensis , *BRUCELLOSIS , *DRUG development , *METHIONYL-tRNA formyltransferase , *PHARMACOKINETICS , *ENZYME inhibitors , *THERAPEUTICS - Abstract
We investigated Brucella melitensis methionyl-tRNA-synthetase (BmMetRS) with molecular, structural and phenotypic methods to learn if BmMetRS is a promising target for brucellosis drug development. Recombinant BmMetRS was expressed, purified from wild type Brucella melitensis biovar Abortus 2308 strain ATCC/CRP #DD-156 and screened by a thermal melt assay against a focused library of one hundred previously classified methionyl-tRNA-synthetase inhibitors of the blood stage form of Trypanosoma brucei. Three compounds showed appreciable shift of denaturation temperature and were selected for further studies on inhibition of the recombinant enzyme activity and cell viability against wild type B. melitensis strain 16M. BmMetRS protein complexed with these three inhibitors resolved into three-dimensional crystal structures and was analyzed. All three selected methionyl-tRNA-synthetase compounds inhibit recombinant BmMetRS enzymatic functions in an aminoacylation assay at varying concentrations. Furthermore, growth inhibition of B. melitensis strain 16M by the compounds was shown. Inhibitor-BmMetRS crystal structure models were used to illustrate the molecular basis of the enzyme inhibition. Our current data suggests that BmMetRS is a promising target for brucellosis drug development. However, further studies are needed to optimize lead compound potency, efficacy and safety as well as determine the pharmacokinetics, optimal dosage, and duration for effective treatment. [ABSTRACT FROM AUTHOR]
- Published
- 2016
- Full Text
- View/download PDF
9. A binding hotspot in Trypanosoma cruzi histidyl-tRNA synthetase revealed by fragment-based crystallographic cocktail screens.
- Author
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Koh, Cho Yeow, Kallur Siddaramaiah, Latha, Ranade, Ranae M., Nguyen, Jasmine, Jian, Tengyue, Zhang, Zhongsheng, Gillespie, J. Robert, Buckner, Frederick S., Verlinde, Christophe L. M. J., Fan, Erkang, and Hol, Wim G. J.
- Subjects
TRYPANOSOMA cruzi ,CHAGAS' disease ,TRANSFER RNA ,SMALL molecules ,TRYPANOSOMATIDAE - Abstract
American trypanosomiasis, commonly known as Chagas disease, is a neglected tropical disease caused by the protozoan parasite Trypanosoma cruzi. The chronic form of the infection often causes debilitating morbidity and mortality. However, the current treatment for the disease is typically inadequate owing to drug toxicity and poor efficacy, necessitating a continual effort to discover and develop new antiparasitic therapeutic agents. The structure of T. cruzi histidyl-tRNA synthetase (HisRS), a validated drug target, has previously been reported. Based on this structure and those of human cytosolic HisRS, opportunities for the development of specific inhibitors were identified. Here, efforts are reported to identify small molecules that bind to T. cruzi HisRS through fragment-based crystallographic screening in order to arrive at chemical starting points for the development of specific inhibitors. T. cruzi HisRS was soaked into 68 different cocktails from the Medical Structural Genomics of Pathogenic Protozoa (MSGPP) fragment library and diffraction data were collected to identify bound fragments after soaking. A total of 15 fragments were identified, all bound to the same site on the protein, revealing a fragment-binding hotspot adjacent to the ATP-binding pocket. On the basis of the initial hits, the design of reactive fragments targeting the hotspot which would be simultaneously covalently linked to a cysteine residue present only in trypanosomatid HisRS was initiated. Inhibition of T. cruzi HisRS was observed with the resultant reactive fragments and the anticipated binding mode was confirmed crystallographically. These results form a platform for the development of future generations of selective inhibitors for trypanosomatid HisRS. [ABSTRACT FROM AUTHOR]
- Published
- 2015
- Full Text
- View/download PDF
10. Synergy Testing of FDA-Approved Drugs Identifies Potent Drug Combinations against Trypanosoma cruzi.
- Author
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Planer, Joseph D., Hulverson, Matthew A., Arif, Jennifer A., Ranade, Ranae M., Don, Robert, and Buckner, Frederick S.
- Subjects
TRYPANOSOMA cruzi ,SEROTONIN uptake inhibitors ,PARACOCCIDIOIDOMYCOSIS ,CHAGAS' disease ,NEGLECTED diseases ,ANTIPARASITIC agents ,CALCIUM antagonists - Abstract
An estimated 8 million persons, mainly in Latin America, are infected with Trypanosoma cruzi, the etiologic agent of Chagas disease. Existing antiparasitic drugs for Chagas disease have significant toxicities and suboptimal effectiveness, hence new therapeutic strategies need to be devised to address this neglected tropical disease. Due to the high research and development costs of bringing new chemical entities to the clinic, we and others have investigated the strategy of repurposing existing drugs for Chagas disease. Screens of FDA-approved drugs (described in this paper) have revealed a variety of chemical classes that have growth inhibitory activity against mammalian stage Trypanosoma cruzi parasites. Aside from azole antifungal drugs that have low or sub-nanomolar activity, most of the active compounds revealed in these screens have effective concentrations causing 50% inhibition (EC
50 's) in the low micromolar or high nanomolar range. For example, we have identified an antihistamine (clemastine, EC50 of 0.4 µM), a selective serotonin reuptake inhibitor (fluoxetine, EC50 of 4.4 µM), and an antifolate drug (pyrimethamine, EC50 of 3.8 µM) and others. When tested alone in the murine model of Trypanosoma cruzi infection, most compounds had insufficient efficacy to lower parasitemia thus we investigated using combinations of compounds for additive or synergistic activity. Twenty-four active compounds were screened in vitro in all possible combinations. Follow up isobologram studies showed at least 8 drug pairs to have synergistic activity on T. cruzi growth. The combination of the calcium channel blocker, amlodipine, plus the antifungal drug, posaconazole, was found to be more effective at lowering parasitemia in mice than either drug alone, as was the combination of clemastine and posaconazole. Using combinations of FDA-approved drugs is a promising strategy for developing new treatments for Chagas disease. Author Summary: Chronic infection with Trypanosoma cruzi causes progressive damage to the heart and other organs that is fatal in about 30% of cases. Known as Chagas disease, this is a major public health problem in Latin America. The existing medicines were developed over forty years ago and are not widely used because of toxicity and unreliable effectiveness. To discover better treatments, we screened a collection of existing drugs for growth inhibitory activity on Trypanosoma cruzi. Several dozen orally administered drugs were discovered, but when used by themselves they were not strong enough to cure the infection in an animal model. We tested a set of 24 of these drugs in every two-way combination and identified eight synergistic partners. At least two of these combinations were able to substantially lower parasite levels in the mouse model of Trypanosoma cruzi infection. Thus, finding pairs of FDA-approved drugs that can be used in combination may be a pragmatic and effective strategy for designing new therapies for Chagas disease. [ABSTRACT FROM AUTHOR]- Published
- 2014
- Full Text
- View/download PDF
11. Synergy Testing of FDA-Approved Drugs Identifies Potent Drug Combinations against Trypanosoma cruzi.
- Author
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Planer, Joseph D., Hulverson, Matthew A., Arif, Jennifer A., Ranade, Ranae M., Don, Robert, and Buckner, Frederick S.
- Subjects
TRYPANOSOMA cruzi ,DRUG additives ,DRUG therapy ,PHARMACOLOGY ,DRUG side effects - Abstract
An estimated 8 million persons, mainly in Latin America, are infected with Trypanosoma cruzi, the etiologic agent of Chagas disease. Existing antiparasitic drugs for Chagas disease have significant toxicities and suboptimal effectiveness, hence new therapeutic strategies need to be devised to address this neglected tropical disease. Due to the high research and development costs of bringing new chemical entities to the clinic, we and others have investigated the strategy of repurposing existing drugs for Chagas disease. Screens of FDA-approved drugs (described in this paper) have revealed a variety of chemical classes that have growth inhibitory activity against mammalian stage Trypanosoma cruzi parasites. Aside from azole antifungal drugs that have low or sub-nanomolar activity, most of the active compounds revealed in these screens have effective concentrations causing 50% inhibition (EC
50 's) in the low micromolar or high nanomolar range. For example, we have identified an antihistamine (clemastine, EC50 of 0.4 µM), a selective serotonin reuptake inhibitor (fluoxetine, EC50 of 4.4 µM), and an antifolate drug (pyrimethamine, EC50 of 3.8 µM) and others. When tested alone in the murine model of Trypanosoma cruzi infection, most compounds had insufficient efficacy to lower parasitemia thus we investigated using combinations of compounds for additive or synergistic activity. Twenty-four active compounds were screened in vitro in all possible combinations. Follow up isobologram studies showed at least 8 drug pairs to have synergistic activity on T. cruzi growth. The combination of the calcium channel blocker, amlodipine, plus the antifungal drug, posaconazole, was found to be more effective at lowering parasitemia in mice than either drug alone, as was the combination of clemastine and posaconazole. Using combinations of FDA-approved drugs is a promising strategy for developing new treatments for Chagas disease. [ABSTRACT FROM AUTHOR]- Published
- 2014
- Full Text
- View/download PDF
12. Structures of Trypanosoma brucei Methionyl-tRNA Synthetase with Urea-Based Inhibitors Provide Guidance for Drug Design against Sleeping Sickness.
- Author
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Koh, Cho Yeow, Kim, Jessica E., Wetzel, Allan B., de van der Schueren, Will J., Shibata, Sayaka, Ranade, Ranae M., Liu, Jiyun, Zhang, Zhongsheng, Gillespie, J. Robert, Buckner, Frederick S., Verlinde, Christophe L. M. J., Fan, Erkang, and Hol, Wim G. J.
- Subjects
DRUG design ,TRYPANOSOMA brucei ,AFRICAN trypanosomiasis ,MULTIENZYME complexes ,DRUG development - Abstract
Methionyl-tRNA synthetase of Trypanosoma brucei (TbMetRS) is an important target in the development of new antitrypanosomal drugs. The enzyme is essential, highly flexible and displaying a large degree of changes in protein domains and binding pockets in the presence of substrate, product and inhibitors. Targeting this protein will benefit from a profound understanding of how its structure adapts to ligand binding. A series of urea-based inhibitors (UBIs) has been developed with IC
50 values as low as 19 nM against the enzyme. The UBIs were shown to be orally available and permeable through the blood-brain barrier, and are therefore candidates for development of drugs for the treatment of late stage human African trypanosomiasis. Here, we expand the structural diversity of inhibitors from the previously reported collection and tested for their inhibitory effect on TbMetRS and on the growth of T. brucei cells. The binding modes and binding pockets of 14 UBIs are revealed by determination of their crystal structures in complex with TbMetRS at resolutions between 2.2 Å to 2.9 Å. The structures show binding of the UBIs through conformational selection, including occupancy of the enlarged methionine pocket and the auxiliary pocket. General principles underlying the affinity of UBIs for TbMetRS are derived from these structures, in particular the optimum way to fill the two binding pockets. The conserved auxiliary pocket might play a role in binding tRNA. In addition, a crystal structure of a ternary TbMetRS•inhibitor•AMPPCP complex indicates that the UBIs are not competing with ATP for binding, instead are interacting with ATP through hydrogen bond. This suggests a possibility that a general 'ATP-engaging' binding mode can be utilized for the design and development of inhibitors targeting tRNA synthetases of other disease-causing pathogen. Author Summary: Infection by the protozoan parasite Trypanosoma brucei causes sleeping sickness, also called human African trypanosomiasis. Without treatment, the disease is fatal yet current therapeutic options are inadequate and better medicines are needed. We have previously reported several potent inhibitors of T. brucei methionyl-tRNA synthetase, an essential enzyme involved in the protein biosynthesis. Recently, a new series of the inhibitors was synthesized which has improved membrane permeability over the earlier inhibitors. When applied to mouse with T. brucei infection, the new compounds are orally available and reach the central nervous system to reduce parasite loads, and therefore are promising molecules to be developed into antitrypanosomal drug. Here, more inhibitors from this series are reported and tested for their activities. High resolution crystal structures were determined that revealed how these inhibitors bind to the target enzyme. The binding pockets of these inhibitors are thoroughly explored, providing profound insights which are beneficial for further development of MetRS inhibitors against sleeping sickness. A ternary complex of the enzyme, an inhibitor, and an ATP analogue was also determined, indicates that the inhibitor does not compete with ATP for binding. Based on this, a general approach to use inhibitors that engage ATP for binding to tRNA synthetases is proposed. [ABSTRACT FROM AUTHOR]- Published
- 2014
- Full Text
- View/download PDF
13. Structures of Trypanosoma brucei Methionyl-tRNA Synthetase with Urea-Based Inhibitors Provide Guidance for Drug Design against Sleeping Sickness.
- Author
-
Koh, Cho Yeow, Kim, Jessica E., Wetzel, Allan B., de van der Schueren, Will J., Shibata, Sayaka, Ranade, Ranae M., Liu, Jiyun, Zhang, Zhongsheng, Gillespie, J. Robert, Buckner, Frederick S., Verlinde, Christophe L. M. J., Fan, Erkang, and Hol, Wim G. J.
- Subjects
TRYPANOSOMA brucei ,METHIONYL transfer RNA ,DRUG design ,TREATMENT of African trypanosomiasis ,LIGAND binding (Biochemistry) - Abstract
Methionyl-tRNA synthetase of Trypanosoma brucei (TbMetRS) is an important target in the development of new antitrypanosomal drugs. The enzyme is essential, highly flexible and displaying a large degree of changes in protein domains and binding pockets in the presence of substrate, product and inhibitors. Targeting this protein will benefit from a profound understanding of how its structure adapts to ligand binding. A series of urea-based inhibitors (UBIs) has been developed with IC
50 values as low as 19 nM against the enzyme. The UBIs were shown to be orally available and permeable through the blood-brain barrier, and are therefore candidates for development of drugs for the treatment of late stage human African trypanosomiasis. Here, we expand the structural diversity of inhibitors from the previously reported collection and tested for their inhibitory effect on TbMetRS and on the growth of T. brucei cells. The binding modes and binding pockets of 14 UBIs are revealed by determination of their crystal structures in complex with TbMetRS at resolutions between 2.2 Å to 2.9 Å. The structures show binding of the UBIs through conformational selection, including occupancy of the enlarged methionine pocket and the auxiliary pocket. General principles underlying the affinity of UBIs for TbMetRS are derived from these structures, in particular the optimum way to fill the two binding pockets. The conserved auxiliary pocket might play a role in binding tRNA. In addition, a crystal structure of a ternary TbMetRS•inhibitor•AMPPCP complex indicates that the UBIs are not competing with ATP for binding, instead are interacting with ATP through hydrogen bond. This suggests a possibility that a general ‘ATP-engaging’ binding mode can be utilized for the design and development of inhibitors targeting tRNA synthetases of other disease-causing pathogen. [ABSTRACT FROM AUTHOR]- Published
- 2014
- Full Text
- View/download PDF
14. Substituted 2-Phenylimidazopyridines:A NewClass of Drug Leads for Human African Trypanosomiasis.
- Author
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Tatipaka, Hari Babu, Gillespie, J. Robert, Chatterjee, Arnab K., Norcross, Neil R., Hulverson, Matthew A., Ranade, Ranae M., Nagendar, Pendem, Creason, Sharon A., McQueen, Joshua, Duster, Nicole A., Nagle, Advait, Supek, Frantisek, Molteni, Valentina, Wenzler, Tanja, Brun, Reto, Glynne, Richard, Buckner, Frederick S., and Gelb, Michael H.
- Published
- 2014
- Full Text
- View/download PDF
15. Urea-Based Inhibitorsof TrypanosomabruceiMethionyl-tRNA Synthetase: Selectivity andin Vivo Characterization.
- Author
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Shibata, Sayaka, Gillespie, J. Robert, Ranade, Ranae M., Koh, Cho Yeow, Kim, Jessica E., Laydbak, Joy U., Zucker, Frank H., Hol, Wim G. J., Verlinde, Christophe L. M. J., Buckner, Frederick S., and Fan, Erkang
- Published
- 2012
- Full Text
- View/download PDF
16. Optimization of a binding fragment targeting the “enlarged methionine pocket” leads to potent Trypanosoma brucei methionyl-tRNA synthetase inhibitors.
- Author
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Huang, Wenlin, Zhang, Zhongsheng, Ranade, Ranae M., Gillespie, J. Robert, Barros-Álvarez, Ximena, Creason, Sharon A., Shibata, Sayaka, Verlinde, Christophe L.M.J., Hol, Wim G.J., Buckner, Frederick S., and Fan, Erkang
- Subjects
- *
ENZYME inhibitors , *METHIONINE , *TRYPANOSOMA brucei , *AMINOACYL-tRNA synthetases , *PROCESS optimization - Abstract
Potent inhibitors of Trypanosoma brucei methionyl-tRNA synthetase were previously designed using a structure-guided approach. Compounds 1 and 2 were the most active compounds in the cyclic and linear linker series, respectively. To further improve cellular potency, SAR investigation of a binding fragment targeting the “enlarged methionine pocket” (EMP) was performed. The optimization led to the identification of a 6,8-dichloro-tetrahydroquinoline ring as a favorable fragment to bind the EMP. Replacement of 3,5-dichloro-benzyl group (the EMP binding fragment) of inhibitor 2 using this tetrahydroquinoline fragment resulted in compound 13 , that exhibited an EC 50 of 4 nM. [ABSTRACT FROM AUTHOR]
- Published
- 2017
- Full Text
- View/download PDF
17. Leishmania donovani tyrosyl-tRNA synthetase structure in complex with a tyrosyl adenylate analog and comparisons with human and protozoan counterparts.
- Author
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Barros-Álvarez, Ximena, Kerchner, Keshia M., Koh, Cho Yeow, Turley, Stewart, Pardon, Els, Steyaert, Jan, Ranade, Ranae M., Gillespie, J. Robert, Zhang, Zhongsheng, Verlinde, Christophe L.M.J., Fan, Erkang, Buckner, Frederick S., and Hol, Wim G.J.
- Subjects
- *
LEISHMANIA donovani , *TRANSFER RNA , *ADENYLATE cyclase , *PROTOZOA , *MONOMERS - Abstract
The crystal structure of Leishmania donovani tyrosyl-tRNA synthetase ( Ld TyrRS) in complex with a nanobody and the tyrosyl adenylate analog TyrSA was determined at 2.75 Å resolution. Nanobodies are the variable domains of camelid heavy chain-only antibodies. The nanobody makes numerous crystal contacts and in addition reduces the flexibility of a loop of Ld TyrRS. TyrSA is engaged in many interactions with active site residues occupying the tyrosine and adenine binding pockets. The Ld TyrRS polypeptide chain consists of two pseudo-monomers, each consisting of two domains. Comparing the two independent chains in the asymmetric unit reveals that the two pseudo-monomers of Ld TyrRS can bend with respect to each other essentially as rigid bodies. This flexibility might be useful in the positioning of tRNA for catalysis since both pseudo-monomers in the Ld TyrRS chain are needed for charging tRNA Tyr . An “extra pocket” (EP) appears to be present near the adenine binding region of Ld TyrRS. Since this pocket is absent in the two human homologous enzymes, the EP provides interesting opportunities for obtaining selective drugs for treating infections caused by L. donovani , a unicellular parasite causing visceral leishmaniasis, or kala azar, which claims 20,000 to 30,000 deaths per year. Sequence and structural comparisons indicate that the EP is a characteristic which also occurs in the active site of several other important pathogenic protozoa. Therefore, the structure of Ld TyrRS could inspire the design of compounds useful for treating several different parasitic diseases. [ABSTRACT FROM AUTHOR]
- Published
- 2017
- Full Text
- View/download PDF
18. Synergy Testing of FDA-Approved Drugs Identifies Potent Drug Combinations against Trypanosoma cruzi.
- Author
-
Planer, Joseph D., Hulverson, Matthew A., Arif, Jennifer A., Ranade, Ranae M., Don, Robert, and Buckner, Frederick S.
- Subjects
- *
TRYPANOSOMA cruzi , *DRUG additives , *DRUG therapy , *PHARMACOLOGY , *DRUG side effects - Abstract
An estimated 8 million persons, mainly in Latin America, are infected with Trypanosoma cruzi, the etiologic agent of Chagas disease. Existing antiparasitic drugs for Chagas disease have significant toxicities and suboptimal effectiveness, hence new therapeutic strategies need to be devised to address this neglected tropical disease. Due to the high research and development costs of bringing new chemical entities to the clinic, we and others have investigated the strategy of repurposing existing drugs for Chagas disease. Screens of FDA-approved drugs (described in this paper) have revealed a variety of chemical classes that have growth inhibitory activity against mammalian stage Trypanosoma cruzi parasites. Aside from azole antifungal drugs that have low or sub-nanomolar activity, most of the active compounds revealed in these screens have effective concentrations causing 50% inhibition (EC50's) in the low micromolar or high nanomolar range. For example, we have identified an antihistamine (clemastine, EC50 of 0.4 µM), a selective serotonin reuptake inhibitor (fluoxetine, EC50 of 4.4 µM), and an antifolate drug (pyrimethamine, EC50 of 3.8 µM) and others. When tested alone in the murine model of Trypanosoma cruzi infection, most compounds had insufficient efficacy to lower parasitemia thus we investigated using combinations of compounds for additive or synergistic activity. Twenty-four active compounds were screened in vitro in all possible combinations. Follow up isobologram studies showed at least 8 drug pairs to have synergistic activity on T. cruzi growth. The combination of the calcium channel blocker, amlodipine, plus the antifungal drug, posaconazole, was found to be more effective at lowering parasitemia in mice than either drug alone, as was the combination of clemastine and posaconazole. Using combinations of FDA-approved drugs is a promising strategy for developing new treatments for Chagas disease. [ABSTRACT FROM AUTHOR]
- Published
- 2014
- Full Text
- View/download PDF
19. Distinct States of Methionyl-tRNA Synthetase Indicate Inhibitor Binding by Conformational Selection
- Author
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Koh, Cho Yeow, Kim, Jessica E., Shibata, Sayaka, Ranade, Ranae M., Yu, Mingyan, Liu, Jiyun, Gillespie, J. Robert, Buckner, Frederick S., Verlinde, Christophe L.M.J., Fan, Erkang, and Hol, Wim G.J.
- Subjects
- *
METHIONYL transfer RNA , *CONFORMATIONAL analysis , *PROTEIN binding , *ENZYME inhibitors , *CRYSTAL structure , *METHIONINE - Abstract
Summary: To guide development of new drugs targeting methionyl-tRNA synthetase (MetRS) for treatment of human African trypanosomiasis, crystal structure determinations of Trypanosoma brucei MetRS in complex with its substrate methionine and its intermediate product methionyl-adenylate were followed by those of the enzyme in complex with high-affinity aminoquinolone inhibitors via soaking experiments. Drastic changes in conformation of one of the two enzymes in the asymmetric unit allowed these inhibitors to occupy an enlarged methionine pocket and a new so-called auxiliary pocket. Interestingly, a small low-affinity compound caused the same conformational changes, removed the methionine without occupying the methionine pocket, and occupied the previously not existing auxiliary pocket. Analysis of these structures indicates that the binding of the inhibitors is the result of conformational selection, not induced fit. [ABSTRACT FROM AUTHOR]
- Published
- 2012
- Full Text
- View/download PDF
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