title: Quantitative structure activity relationship of IA3-like peptides as aspartic proteinase inhibitors.
abstract: The IA(3) polypeptide inhibitor from Saccharomyces cerevisiae interacts potently and selectively with its target, the S. cerevisiae vacuolar aspartic proteinase (ScPr). Upon encountering the enzyme, residues 2-32 of the intrinsically unstructured IA(3) polypeptide become ordered into an almost-perfect alpha-helix. In previous IA(3) mutagenesis studies, we identified important characteristics of the enzyme inhibitor interactions and generated a large dataset of variants with K(i) values determined experimentally at pH 3.1 and 4.7. Using this information, the three-dimensional structure of each variant was modelled in silico with the correct protonation for each experimental pH value. A set of descriptors of the inhibitor/ScPr interactions was then calculated and used to establish mathematical models relating the variant sequences to their inhibitory activities at each pH. Cross-validation, external-set validation and five separate selections of the training and test samples confirmed the robustness of the equations. A major contributor to the structure-activity relationship was the free energy of binding calculated by the FoldX program. The mathematical models were challenged further (i) by in silico alanine-scanning mutagenesis of residues 2-32 in IA(3) and relating binding energy to experimentally derived inhibition constants for selected representatives of these variants; and (ii) by predicting inhibitory-potencies for two novel IA(3)-variants. The predictions of the equations for these new IA(3)-variants with ScPr matched almost precisely the kinetic data determined experimentally. The models described represent valuable tools for the future design of novel inhibitor variants active against ScPr and other aspartic proteinases.
authors.affiliation: Laboratory of Theoretical and Computational Chemistry, Chemistry Faculty, Havana University, 10400, Havana, Cuba.
authors.forename: Juan Alexander
authors.initials: JA
authors.lastname: Padrón-García
authors.forename: Manuel
authors.initials: M
authors.lastname: Alonso-Tarajano
authors.forename: Esther
authors.initials: E
authors.lastname: Alonso-Becerra
authors.forename: Tim J
authors.initials: TJ
authors.lastname: Winterburn
authors.forename: Yasser
authors.initials: Y
authors.lastname: Ruiz
authors.forename: John
authors.initials: J
authors.lastname: Kay
authors.forename: Colin
authors.initials: C
authors.lastname: Berry
mesh_terms: D000409:Alanine / Q000378:metabolism; D000595:Amino Acid Sequence; D019943:Amino Acid Substitution; D016282:Aspartic Acid Endopeptidases / Q000037:antagonists & inhibitors* / Q000737:chemistry* / Q000378:metabolism; D003198:Computer Simulation; D004926:Escherichia coli / Q000235:genetics; D006863:Hydrogen-Ion Concentration; D008958:Models, Molecular; D008969:Molecular Sequence Data; D009154:Mutation; D021281:Quantitative Structure-Activity Relationship; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D015203:Reproducibility of Results; D012441:Saccharomyces cerevisiae / Q000201:enzymology / Q000378:metabolism; D029701:Saccharomyces cerevisiae Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D013816:Thermodynamics
chemical_list: C018134:PAI3 protein, S cerevisiae; D011994:Recombinant Proteins; D029701:Saccharomyces cerevisiae Proteins; D016282:Aspartic Acid Endopeptidases; D000409:Alanine