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dc.contributor.authorLin, Kuan-Hung
dc.contributor.authorNalivaika, Ellen A.
dc.contributor.authorPrachanronarong, Kristina L.
dc.contributor.authorYilmaz, Nese Kurt
dc.contributor.authorSchiffer, Celia A.
dc.date2022-08-11T08:10:52.000
dc.date.accessioned2022-08-23T17:23:01Z
dc.date.available2022-08-23T17:23:01Z
dc.date.issued2016-10-14
dc.date.submitted2017-02-17
dc.identifier.citationACS Infect Dis. 2016 Oct 14;2(10):734-743. Epub 2016 Sep 22. <a href="https://doi.org/10.1021/acsinfecdis.6b00131">Link to article on publisher's site</a>
dc.identifier.issn2373-8227 (Linking)
dc.identifier.doi10.1021/acsinfecdis.6b00131
dc.identifier.pmid27657335
dc.identifier.urihttp://hdl.handle.net/20.500.14038/48868
dc.description.abstractDengue virus (DENV), transmitted predominantly in tropical and subtropical regions by the mosquito Aedes aegypti, infects millions of people and leads to dengue fever and thousands of deaths each year. There are no direct-acting antivirals to combat DENV, and molecular and structural knowledge is required to develop such compounds. The dengue NS2B/NS3 protease is a promising target for direct-acting antivirals, as viral polyprotein cleavage during replication is required for the maturation of the viral particle. The NS2B/NS3 protease processes 8 of the 13 viral polyprotein cleavage sites to allow viral maturation. Although these sites share little sequence homology beyond the P1 and P2 positions, most are well conserved among the serotypes. How the other substrate residues, especially at the P' side, affect substrate recognition remains unclear. We exploited the tight-binding general serine protease inhibitor aprotinin to investigate protease-substrate interactions at the molecular level. We engineered aprotinin's binding loop with sequences mimicking the P' side of DENV substrates. P' residues significantly modulate substrate affinity to protease, with inhibition constants varying from nanomolar to sub-millimolar. Structural and dynamic analysis revealed the molecular basis of this modulation and allowed identifying optimal residues for each of the P' positions. In addition, isothermal titration calorimetry showed binding to be solely entropy driven for all constructs. Potential flaviviral P' side inhibitors could benefit from mimicking the optimal residues at P' positions and incorporate hydrophobicity and rigidity to maintain entropic advantage for potency.
dc.language.isoen_US
dc.relation<a href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=pubmed&cmd=Retrieve&list_uids=27657335&dopt=Abstract">Link to Article in PubMed</a>
dc.relation.urlhttps://doi.org/10.1021/acsinfecdis.6b00131
dc.subjectdengue
dc.subjectpeptide sequence
dc.subjectprotease inhibitor
dc.subjectsubstrate recognition
dc.subjectviral protease
dc.subjectBiochemistry
dc.subjectEnzymes and Coenzymes
dc.subjectMedicinal Chemistry and Pharmaceutics
dc.subjectMedicinal-Pharmaceutical Chemistry
dc.subjectMolecular Biology
dc.subjectStructural Biology
dc.subjectVirus Diseases
dc.titleDengue Protease Substrate Recognition: Binding of the Prime Side
dc.typeJournal Article
dc.source.journaltitleACS infectious diseases
dc.source.volume2
dc.source.issue10
dc.identifier.legacycoverpagehttps://escholarship.umassmed.edu/schiffer/15
dc.identifier.contextkey9705344
html.description.abstract<p>Dengue virus (DENV), transmitted predominantly in tropical and subtropical regions by the mosquito Aedes aegypti, infects millions of people and leads to dengue fever and thousands of deaths each year. There are no direct-acting antivirals to combat DENV, and molecular and structural knowledge is required to develop such compounds. The dengue NS2B/NS3 protease is a promising target for direct-acting antivirals, as viral polyprotein cleavage during replication is required for the maturation of the viral particle. The NS2B/NS3 protease processes 8 of the 13 viral polyprotein cleavage sites to allow viral maturation. Although these sites share little sequence homology beyond the P1 and P2 positions, most are well conserved among the serotypes. How the other substrate residues, especially at the P' side, affect substrate recognition remains unclear. We exploited the tight-binding general serine protease inhibitor aprotinin to investigate protease-substrate interactions at the molecular level. We engineered aprotinin's binding loop with sequences mimicking the P' side of DENV substrates. P' residues significantly modulate substrate affinity to protease, with inhibition constants varying from nanomolar to sub-millimolar. Structural and dynamic analysis revealed the molecular basis of this modulation and allowed identifying optimal residues for each of the P' positions. In addition, isothermal titration calorimetry showed binding to be solely entropy driven for all constructs. Potential flaviviral P' side inhibitors could benefit from mimicking the optimal residues at P' positions and incorporate hydrophobicity and rigidity to maintain entropic advantage for potency.</p>
dc.identifier.submissionpathschiffer/15
dc.contributor.departmentDepartment of Biochemistry and Molecular Pharmacology
dc.source.pages734-743


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