Mutational and Structural Analysis of Conserved Residues in Ribose-5-Phosphate Isomerase B from Leishmania donovani: Role in Substrate Recognition and Conformational Stability

Ribose-5-phosphate isomerase B from Leishmania donovani (LdRpiB) is one of the potential drug targets against visceral leishmaniasis. In the present study, we have targeted several conserved amino acids for mutational analysis (i.e. Cys69, His11, His102, His138, Asp45, Tyr46, Pro47 and Glu149) to ga...

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Published in:PloS one 2016-03, Vol.11 (3), p.e0150764-e0150764
Main Authors: Kaur, Preet Kamal, Tripathi, Neha, Desale, Jayesh, Neelagiri, Soumya, Yadav, Shailendra, Bharatam, Prasad V, Singh, Sushma
Format: Article
Language:English
Subjects:
Aldose-Ketose Isomerases - chemistry
Aldose-Ketose Isomerases - genetics
Aldose-Ketose Isomerases - metabolism
Amino Acid Sequence
Amino Acid Substitution
Amino acids
Analysis
Arabinose
Atomic structure
Binding
Biology and Life Sciences
Biotechnology
Catalysis
Clostridium difficile
Clostridium thermocellum
Conserved Sequence
D-Ribose
Data processing
Drug development
E coli
Enzymatic activity
Enzyme activity
Enzymes
Fluorescence
Health education
Kinetics
Leishmania donovani
Leishmania donovani - genetics
Leishmania donovani - metabolism
Leishmaniasis
Models, Molecular
Molecular Docking Simulation
Molecular Dynamics Simulation
Molecular Sequence Data
Mutagenesis
Mutants
Mutation
Mycobacterium tuberculosis
Parasites
Parasitic diseases
Pharmaceutical sciences
Phosphates
Physical Sciences
Protein Binding
Protein Conformation
Protein structure
Proteins
Ribose
Ribose-5-phosphate isomerase
Risk factors
Sequence Alignment
Spectral analysis
Stability
Structural analysis
Structure-Activity Relationship
Substrate Specificity
Substrates
Trypanosoma cruzi
Tuberculosis
Vector-borne diseases
Visceral leishmaniasis
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cited_by cdi_FETCH-LOGICAL-c692t-483fbe3ff0039db535a88d4f5dacf92e4f6e6cd776a078329012833fe4106ce43
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Tripathi, Neha
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Singh, Sushma
description Ribose-5-phosphate isomerase B from Leishmania donovani (LdRpiB) is one of the potential drug targets against visceral leishmaniasis. In the present study, we have targeted several conserved amino acids for mutational analysis (i.e. Cys69, His11, His102, His138, Asp45, Tyr46, Pro47 and Glu149) to gain crucial insights into their role in substrate binding, catalysis and conformational stability of the enzyme. All the eight LdRpiB variants were cloned, sequenced, expressed and purified. C69S, H102N, D45N and E149A mutants exhibited complete loss of enzyme activity indicating that they are indispensable for the enzyme activity. Kinetic parameters were altered in case of H138N, H11N and P47A variants; however Y46F exhibited similar kinetic behaviour as wild type. All the mutants except H138N exhibited altered protein structure as determined by CD and fluorescence spectral analysis. This data was supported by the atomic level details of the conformational changes and substrate binding using molecular dynamic simulations. LdRpiB also exhibited activity with D-form of various aldose substrates in the order of D-ribose > D-talose > D-allose > D-arabinose. Our study provides insights for better understanding of substrate enzyme interactions which can rationalize the process of drug design against parasite RpiB.
doi_str_mv 10.1371/journal.pone.0150764
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Academic</collection><collection>PubMed Central (Full Participant titles)</collection><collection>DOAJ Directory of Open Access Journals</collection><jtitle>PloS one</jtitle></facets><delivery><delcategory>Remote Search Resource</delcategory><fulltext>fulltext</fulltext></delivery><addata><au>Kaur, Preet Kamal</au><au>Tripathi, Neha</au><au>Desale, Jayesh</au><au>Neelagiri, Soumya</au><au>Yadav, Shailendra</au><au>Bharatam, Prasad V</au><au>Singh, Sushma</au><au>Salsbury, Freddie</au><format>journal</format><genre>article</genre><ristype>JOUR</ristype><atitle>Mutational and Structural Analysis of Conserved Residues in Ribose-5-Phosphate Isomerase B from Leishmania donovani: Role in Substrate Recognition and Conformational Stability</atitle><jtitle>PloS one</jtitle><addtitle>PLoS One</addtitle><date>2016-03-08</date><risdate>2016</risdate><volume>11</volume><issue>3</issue><spage>e0150764</spage><epage>e0150764</epage><pages>e0150764-e0150764</pages><issn>1932-6203</issn><eissn>1932-6203</eissn><abstract>Ribose-5-phosphate isomerase B from Leishmania donovani (LdRpiB) is one of the potential drug targets against visceral leishmaniasis. In the present study, we have targeted several conserved amino acids for mutational analysis (i.e. Cys69, His11, His102, His138, Asp45, Tyr46, Pro47 and Glu149) to gain crucial insights into their role in substrate binding, catalysis and conformational stability of the enzyme. All the eight LdRpiB variants were cloned, sequenced, expressed and purified. C69S, H102N, D45N and E149A mutants exhibited complete loss of enzyme activity indicating that they are indispensable for the enzyme activity. Kinetic parameters were altered in case of H138N, H11N and P47A variants; however Y46F exhibited similar kinetic behaviour as wild type. All the mutants except H138N exhibited altered protein structure as determined by CD and fluorescence spectral analysis. This data was supported by the atomic level details of the conformational changes and substrate binding using molecular dynamic simulations. LdRpiB also exhibited activity with D-form of various aldose substrates in the order of D-ribose &gt; D-talose &gt; D-allose &gt; D-arabinose. Our study provides insights for better understanding of substrate enzyme interactions which can rationalize the process of drug design against parasite RpiB.</abstract><cop>United States</cop><pub>Public Library of Science</pub><pmid>26953696</pmid><doi>10.1371/journal.pone.0150764</doi><oa>free_for_read</oa></addata></record>
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identifier ISSN: 1932-6203
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issn 1932-6203
1932-6203
language eng
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source Open Access: PubMed Central; Publicly Available Content Database
subjects Aldose-Ketose Isomerases - chemistry
Aldose-Ketose Isomerases - genetics
Aldose-Ketose Isomerases - metabolism
Amino Acid Sequence
Amino Acid Substitution
Amino acids
Analysis
Arabinose
Atomic structure
Binding
Biology and Life Sciences
Biotechnology
Catalysis
Clostridium difficile
Clostridium thermocellum
Conserved Sequence
D-Ribose
Data processing
Drug development
E coli
Enzymatic activity
Enzyme activity
Enzymes
Fluorescence
Health education
Kinetics
Leishmania donovani
Leishmania donovani - genetics
Leishmania donovani - metabolism
Leishmaniasis
Models, Molecular
Molecular Docking Simulation
Molecular Dynamics Simulation
Molecular Sequence Data
Mutagenesis
Mutants
Mutation
Mycobacterium tuberculosis
Parasites
Parasitic diseases
Pharmaceutical sciences
Phosphates
Physical Sciences
Protein Binding
Protein Conformation
Protein structure
Proteins
Ribose
Ribose-5-phosphate isomerase
Risk factors
Sequence Alignment
Spectral analysis
Stability
Structural analysis
Structure-Activity Relationship
Substrate Specificity
Substrates
Trypanosoma cruzi
Tuberculosis
Vector-borne diseases
Visceral leishmaniasis
title Mutational and Structural Analysis of Conserved Residues in Ribose-5-Phosphate Isomerase B from Leishmania donovani: Role in Substrate Recognition and Conformational Stability
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