title: Learning from directed evolution: theoretical investigations into cooperative mutations in lipase enantioselectivity.
abstract: Molecular modeling with classical force-fields has been used to study the reactant complex and the tetrahedral intermediate in lipase-catalyzed ester hydrolysis in 20 enzyme/substrate combinations. The R and S enantiomers of alpha-methyldecanoic acid ester served as substrates for the wild-type lipase from Pseudomonas aeruginosa and nine selected mutants. After suitable preparation of initial structures from an available wild-type crystal structure, each system was subjected to 1 ns CHARMM force-field molecular dynamics simulations. The resulting geometric and energetic changes allow interpretation of some experimentally observed effects of mutations, particularly with regard to the "hot spots" at residues 155 and 162. The replacement S155F enhances S enantiopreference through a steric relay involving Leu162. The double mutation S53P + L162G improves S enantioselectivity by creating a new binding pocket for the S enantiomer with an additional stabilizing hydrogen bond to His83. The simulations provide insight into remote and cooperative effects of mutations.
authors.affiliation: Max-Planck-Institut für Kohlenforschung, Kaiser-Wilhelm-Platz 1, 45470 Mülheim an der Ruhr, Germany.
authors.forename: Marco
authors.initials: M
authors.lastname: Bocola
authors.forename: Nikolaj
authors.initials: N
authors.lastname: Otte
authors.forename: Karl-Erich
authors.initials: KE
authors.lastname: Jaeger
authors.forename: Manfred T
authors.initials: MT
authors.lastname: Reetz
authors.forename: Walter
authors.initials: W
authors.lastname: Thiel
mesh_terms: D002384:Catalysis; D003198:Computer Simulation; D018360:Crystallography, X-Ray; D019020:Directed Molecular Evolution; D004952:Esters / Q000378:metabolism; D006868:Hydrolysis; D007700:Kinetics; D008049:Lipase / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism*; D008958:Models, Molecular; D015394:Molecular Structure; D009154:Mutation / Q000235:genetics*; D017434:Protein Structure, Tertiary; D011550:Pseudomonas aeruginosa / Q000737:chemistry / Q000201:enzymology / Q000235:genetics; D013237:Stereoisomerism; D013379:Substrate Specificity
chemical_list: D004952:Esters; D008049:Lipase