scholarly journals Molecular mechanism of a parasite kinesin motor and implications for its inhibition

2021 ◽  
Author(s):  
Alexander D. Cook ◽  
Anthony Roberts ◽  
Joseph Atherton ◽  
Rita Tewari ◽  
Maya Topf ◽  
...  
Keyword(s):  
Bound States ◽  
Sequence Divergence ◽  
Selective Inhibition ◽  
Biochemical Properties ◽  
Drug Binding ◽  
Motor Domain ◽  
Complex Life Cycle ◽  
Drug Binding Site ◽  
Current Therapies ◽  
Directed Motility

ABSTRACTPlasmodium parasites cause malaria and are responsible annually for hundreds of thousands of deaths. They have a complex life cycle in which distinct stages are transmitted between, and reproduce in, human and mosquito hosts. In the light of emerging resistance to current therapies, components of the parasite replicative machinery are potentially important targets for anti-parasite drugs. Members of the superfamily of kinesin motors play important roles in the microtubule-based replicative spindle machinery, and kinesin-5 motors are established anti-mitotic targets in other disease contexts. We therefore studied kinesin-5 from Plasmodium falciparum (PfK5) and characterised the biochemical properties and structure of the PfK5 motor domain. We found that the PfK5 motor domain is an ATPase with microtubule plus-end directed motility. We used cryo-EM to determine the motor’s microtubule-bound structure in no nucleotide and AMPPNP-bound states. Despite significant sequence divergence in this motor, these structures reveal that this parasite motor exhibits classical kinesin mechanochemistry. This includes ATP-induced neck-linker docking to the motor domain, which is consistent with the motor’s plus-ended directed motility. Crucially, we also observed that a large insertion in loop5 of the PfK5 motor domain creates a dramatically different chemical environment in the well characterised human kinesin-5 drug-binding site. Our data thereby reveal the possibility for selective inhibition of PfK5 and can be used to inform future exploration of Plasmodium kinesins as anti-parasite targets.

Drug Repurposing Studies Targeting SARS-nCoV2: An Ensemble Docking Approach on Drug Target 3C-like Protease (3CLpro)

2020 ◽  
Author(s):  
Shruti Koulgi ◽  
Vinod Jani ◽  
Mallikarjunachari Uppuladinne ◽  
Uddhavesh Sonavane ◽  
Asheet Kumar Nath ◽  
...  
Keyword(s):  
Drug Repurposing ◽  
Drug Binding ◽  
Ensemble Docking ◽  
Drug Molecules ◽  
Drug Binding Site ◽  
Main Protease ◽  
Docking Approach ◽  
Novel Coronavirus ◽  
Markov State Modeling ◽  
Viral Polyprotein

<p>The COVID-19 pandemic has been responsible for several deaths worldwide. The causative agent behind this disease is the Severe Acute Respiratory Syndrome – novel Coronavirus 2 (SARS-nCoV2). SARS-nCoV2 belongs to the category of RNA viruses. The main protease, responsible for the cleavage of the viral polyprotein is considered as one of the hot targets for treating COVID-19. Earlier reports suggest the use of HIV anti-viral drugs for targeting the main protease of SARS-CoV, which caused SARS in the year 2002-03. Hence, drug repurposing approach may prove to be useful in targeting the main protease of SARS-nCoV2. The high-resolution crystal structure of 3CL<sup>pro</sup> (main protease) of SARS-nCoV2 (PDB ID: 6LU7) was used as the target. The Food and Drug Administration (FDA) approved and SWEETLEAD database of drug molecules were screened. The apo form of the main protease was simulated for a cumulative of 150 ns and 10 μs open source simulation data was used, to obtain conformations for ensemble docking. The representative structures for docking were selected using RMSD-based clustering and Markov State Modeling analysis. This ensemble docking approach for main protease helped in exploring the conformational variation in the drug binding site of the main protease leading to efficient binding of more relevant drug molecules. The drugs obtained as best hits from the ensemble docking possessed anti-bacterial and anti-viral properties. Small molecules with these properties may prove to be useful to treat symptoms exhibited in COVID-19. This <i>in-silico</i> ensemble docking approach would support identification of potential candidates for repurposing against COVID-19.</p>

scholarly journals Plasmodium falciparum DHODH inhibitor complexes reveal flexible binding site

2014 ◽  
Vol 70 (a1) ◽  
pp. C1793-C1793
Author(s):  
Paul Rowland ◽  
Onkar SINGH ◽  
Leila Ross ◽  
Francisco Gamo ◽  
Maria Lafuente-Monasterio ◽  
...  
Keyword(s):  
Drug Resistance ◽  
Binding Site ◽  
Crystal Structures ◽  
Point Mutations ◽  
Crystal Form ◽  
Drug Binding ◽  
Binding Modes ◽  
Resistance Mutations ◽  
Drug Binding Site

Malaria is a preventable and treatable disease, yet annually there are still hundreds of thousands of malaria-related deaths. The disease is caused by infection with mosquito-borne Plasmodium parasites. With hundreds of millions of cases each year there is a very high potential for drug resistance and this has compromised many existing therapies. One target under investigation is the enzyme dihydroorotate dehydrogenase (DHODH) which catalyses the rate-limiting step of pyrimidine biosynthesis and is an essential enzyme in the malaria parasite. There are currently several Plasmodium-selective DHODH inhibitors under development. To investigate the potential for drug resistance against DHODH inhibitors in vitro resistance selections were carried out using known inhibitors from different structural classes [1]. These studies identified point mutations in the drug binding site which lead to reduced sensitivity to the inhibitors, and in some cases increased sensitivity to a different inhibitor, suggesting a novel combination therapy approach to combat resistance. To help understand the significance of the inhibitor binding site mutations we determined the crystal structures of P. falciparum DHODH in complex with the inhibitors Genz-669178, IDI-6253 and IDI-6273. Co-crystallisation experiments led to a new crystal form in each case. Here we describe the crystal structures, the binding modes of the inhibitors and the great flexibility of the binding site, which is able to adjust to accommodate different inhibitor series. The structural role of the resistance mutations is also discussed.

scholarly journals All-Atomic Molecular Dynamic Studies of Human and Drosophila CDK8: Insights into Their Kinase Domains, the LXXLL Motifs, and Drug Binding Site

2020 ◽  
Vol 21 (20) ◽  
pp. 7511
Author(s):  
Wu Xu ◽  
Xiao-Jun Xie ◽  
Ali K. Faust ◽  
Mengmeng Liu ◽  
Xiao Li ◽  
...  
Keyword(s):  
Hydrogen Bond ◽  
Hydrogen Bonds ◽  
Binding Site ◽  
Molecular Dynamic ◽  
Kinase Activity ◽  
Structural Difference ◽  
Drug Binding ◽  
Local Structures ◽  
Drug Binding Site ◽  
Lxxll Motif

Cyclin-dependent kinase 8 (CDK8) and its regulatory partner Cyclin C (CycC) play conserved roles in modulating RNA polymerase II (Pol II)-dependent gene expression. To understand the structure and function relations of CDK8, we analyzed the structures of human and Drosophila CDK8 proteins using molecular dynamics simulations, combined with functional analyses in Drosophila. Specifically, we evaluated the structural differences between hCDK8 and dCDK8 to predict the effects of the LXXLL motif mutation (AQKAA), the P154L mutations, and drug binding on local structures of the CDK8 proteins. First, we have observed that both the LXXLL motif and the kinase activity of CDK8 are required for the normal larval-to-pupal transition in Drosophila. Second, our molecular dynamic analyses have revealed that hCDK8 has higher hydrogen bond occupation of His149-Asp151 and Asp151-Asn156 than dCDK8. Third, the substructure of Asp282, Phe283, Arg285, Thr287 and Cys291 can distinguish human and Drosophila CDK8 structures. In addition, there are two hydrogen bonds in the LXXLL motif: a lower occupation between L312 and L315, and a relatively higher occupation between L312 and L316. Human CDK8 has higher hydrogen bond occupation between L312 and L316 than dCDK8. Moreover, L312, L315 and L316 in the LXXLL motif of CDK8 have the specific pattern of hydrogen bonds and geometries, which could be crucial for the binding to nuclear receptors. Furthermore, the P154L mutation dramatically decreases the hydrogen bond between L312 and L315 in hCDK8, but not in dCDK8. The mutations of P154L and AQKAA modestly alter the local structures around residues 154. Finally, we identified the inhibitor-induced conformational changes of hCDK8, and our results suggest a structural difference in the drug-binding site between hCDK8 and dCDK8. Taken together, these results provide the structural insights into the roles of the LXXLL motif and the kinase activity of CDK8 in vivo.

scholarly journals Structural Plasticity and Distinct Drug-Binding Modes of LfrR, a Mycobacterial Efflux Pump Regulator

2009 ◽  
Vol 191 (24) ◽  
pp. 7531-7537 ◽  
Author(s):  
Marco Bellinzoni ◽  
Silvia Buroni ◽  
Francis Schaeffer ◽  
Giovanna Riccardi ◽  
Edda De Rossi ◽  
...  
Keyword(s):  
Mycobacterium Smegmatis ◽  
Efflux Pump ◽  
Drug Binding ◽  
Binding Modes ◽  
Transcriptional Repressors ◽  
Gene Encoding ◽  
Drug Binding Site ◽  
Conformational Plasticity ◽  
Local Unfolding ◽  
Calorimetric Studies

ABSTRACT The TetR-like transcriptional repressor LfrR controls the expression of the gene encoding the Mycobacterium smegmatis efflux pump LfrA, which actively extrudes fluoroquinolones, cationic dyes, and anthracyclines from the cell and promotes intrinsic antibiotic resistance. The crystal structure of the apoprotein form of the repressor reveals a structurally asymmetric homodimer exhibiting local unfolding and a blocked drug-binding site, emphasizing the significant conformational plasticity of the protein necessary for DNA and multidrug recognition. Crystallographic and calorimetric studies of LfrR-drug complexes further confirm the intrinsic flexibility of the homodimer, which provides a dynamic mechanism to broaden multidrug binding specificity and may be a general property of transcriptional repressors regulating microbial efflux pump expression.

scholarly journals Mutations in Plasmodium falciparumCytochrome b That Are Associated with Atovaquone Resistance Are Located at a Putative Drug-Binding Site

2000 ◽  
Vol 44 (8) ◽  
pp. 2100-2108 ◽  
Author(s):  
Michael Korsinczky ◽  
Nanhua Chen ◽  
Barbara Kotecka ◽  
Allan Saul ◽  
Karl Rieckmann ◽  
...  
Keyword(s):  
Amino Acid ◽  
Binding Site ◽  
Point Mutations ◽  
Drug Binding ◽  
Single Amino Acid ◽  
Malaria Parasites ◽  
Drug Binding Site ◽  
Sole Agent ◽  
Amino Acid Mutations

ABSTRACT Atovaquone is the major active component of the new antimalarial drug Malarone. Considerable evidence suggests that malaria parasites become resistant to atovaquone quickly if atovaquone is used as a sole agent. The mechanism by which the parasite develops resistance to atovaquone is not yet fully understood. Atovaquone has been shown to inhibit the cytochrome bc 1 (CYTbc 1) complex of the electron transport chain of malaria parasites. Here we report point mutations in Plasmodium falciparum CYT b that are associated with atovaquone resistance. Single or double amino acid mutations were detected from parasites that originated from a cloned line and survived various concentrations of atovaquone in vitro. A single amino acid mutation was detected in parasites isolated from a recrudescent patient following atovaquone treatment. These mutations are associated with a 25- to 9,354-fold range reduction in parasite susceptibility to atovaquone. Molecular modeling showed that amino acid mutations associated with atovaquone resistance are clustered around a putative atovaquone-binding site. Mutations in these positions are consistent with a reduced binding affinity of atovaquone for malaria parasite CYTb.

A rapid-response fluorescent probe for the sensitive and selective detection of human albumin in plasma and cell culture supernatants

2016 ◽  
Vol 52 (36) ◽  
pp. 6064-6067 ◽  
Author(s):  
Yi-Ru Wang ◽  
Lei Feng ◽  
Liang Xu ◽  
Yan Li ◽  
Dan-Dan Wang ◽  
...  
Keyword(s):  
Cell Culture ◽  
Binding Site ◽  
Fluorescent Probe ◽  
Human Albumin ◽  
Drug Binding ◽  
Rapid Response ◽  
Sensitive Detection ◽  
Selective Detection ◽  
Drug Binding Site ◽  
Culture Supernatants

A rapid-response fluorescent probeACDMwas developed for selective and sensitive detection of human albumin (HA)viabinding on a non-drug binding site.

scholarly journals Spectroscopic characterization of the warfarin drug-binding site of folded and unfolded human serum albumin anchored on gold nanoparticles: effect of bioconjugation on the loading capacity

RSC Advances ◽  
2018 ◽  
Vol 8 (14) ◽  
pp. 7523-7532 ◽  
Author(s):  
Saba A. J. Sulaiman ◽  
Tanujjal Bora ◽  
Osama K. Abou-Zied
Keyword(s):  
Gold Nanoparticles ◽  
Steady State ◽  
Ultrafast Spectroscopy ◽  
Drug Loading ◽  
Spectroscopic Characterization ◽  
Binding Activity ◽  
Drug Binding ◽  
Loading Capacity ◽  
Drug Binding Site

This work investigates the steady-state and ultrafast spectroscopy of bioconjugated gold nanoparticles and the implications on the protein binding activity and drug-loading capacity.

Quinoline resistance mechanisms in Plasmodium falciparum: the debate goes on

Parasitology ◽  
1997 ◽  
Vol 114 (7) ◽  
pp. 125-136 ◽  
Author(s):  
S. A. WARD ◽  
P. G. BRAY ◽  
S. R. HAWLEY
Keyword(s):  
Efflux Pump ◽  
Resistance Mechanism ◽  
Resistance Mechanisms ◽  
Food Vacuole ◽  
Drug Binding ◽  
Drug Uptake ◽  
Cross Resistance ◽  
Parasite Resistance ◽  
Therapeutic Success ◽  
Drug Binding Site

Despite considerable therapeutic success with the antimalarial 4-aminoquinolines such as chloroquine, there is serious doubt about the future of this drug class due mainly to the development and spread of parasite resistance throughout endemic areas. In this article we review the possible biochemical and molecular basis of resistance. Based on our current understanding we have considered the possibility of developing strategies which may allow the aminoquinolines to once again be used effectively against P. falciparum. Our conclusions are that drug resistance is the result of a reduced rate of drug uptake which in turn reduces the amount of drug available to bind the target. The basis for this reduced accumulation could be an altered pH gradient making the food vacuole more alkaline or the parasite cytosol more acidic, an efflux pump removing drug directly from the membrane or any other process which will reduce the rate of drug uptake. Central to the effectiveness of this resistance mechanism is the transient availability of a high affinity, low capacity drug binding site (possibly haem) within the parasite. Resistance reversers such as verapamil influence the apparent Ka for this drug binding phenomenon via an increased drug uptake rate. We demonstrate that by chemical modification of the aminoquinolines, producing predictable alterations in their physicochemical properties, that it is possible to minimise the verapamil sensitive component of resistance and reduce significantly cross-resistance patterns without loss in absolute activity. Based on these views we suggest that the aminoquinoline antimalarials still have a role to play in the cheap, safe and effective chemotherapy of falciparum malaria.

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