title: Allosteric modulation of caspase 3 through mutagenesis.
abstract: A mutation in the allosteric site of the caspase 3 dimer interface of Val266 to histidine abolishes activity of the enzyme, and models predict that the mutation mimics the action of small molecule allosteric inhibitors by preventing formation of the active site. Mutations were coupled to His266 at two sites in the interface, E124A and Y197C. We present results from X-ray crystallography, enzymatic activity and molecular dynamics simulations for seven proteins, consisting of single, double and triple mutants. The results demonstrate that considering allosteric inhibition of caspase 3 as a shift between discrete 'off-state' or 'on-state' conformations is insufficient. Although His266 is accommodated in the interface, the structural defects are propagated to the active site through a helix on the protein surface. A more comprehensive view of allosteric regulation of caspase 3 requires the representation of an ensemble of inactive states and shows that subtle structural changes lead to the population of the inactive ensemble.
authors.affiliation: Department of Molecular and Structural Biochemistry, North Carolina State University, Raleigh, NC 27695, USA.
authors.forename: Jad
authors.initials: J
authors.lastname: Walters
authors.forename: Joshua L
authors.initials: JL
authors.lastname: Schipper
authors.forename: Paul
authors.initials: P
authors.lastname: Swartz
authors.forename: Carla
authors.initials: C
authors.lastname: Mattos
authors.forename: A Clay
authors.initials: AC
authors.lastname: Clark
mesh_terms: D000494:Allosteric Regulation / Q000235:genetics; D000495:Allosteric Site / Q000235:genetics*; D019943:Amino Acid Substitution; D053148:Caspase 3 / Q000737:chemistry* / Q000235:genetics; D020134:Catalytic Domain; D018360:Crystallography, X-Ray; D006801:Humans; D006860:Hydrogen Bonding; D056004:Molecular Dynamics Simulation; D016297:Mutagenesis, Site-Directed; D054730:Protein Interaction Domains and Motifs; D017433:Protein Structure, Secondary
chemical_list: C505407:CASP3 protein, human; D053148:Caspase 3