title: Relative binding free energy calculations of inhibitors to two mutants (Glu46----Ala/Gln) of ribonuclease T1 using molecular dynamics/free energy perturbation approaches.
abstract: We present free energy perturbation calculations on the complexes of Glu46----Ala46 (E46A) and Glu46----Gln46 (E46Q) mutants of ribonuclease T1 (RNaseT1) with inhibitors 2'-guanosine monophosphate (GMP) and 2'-adenosine monophosphate (AMP) by a thermodynamic perturbation method implemented with molecular dynamics (MD). Using the available crystal structure of the RNaseT1-GMP complex, the structures of E46A-GMP and E46Q-GMP were model built and equilibrated with MD simulations. The structures of E46A-AMP and E46Q-AMP were obtained as a final structure of the GMP----AMP perturbation calculation respectively. The calculated difference in the free energy of binding (delta delta Gbind) was 0.31 kcal/mol for the E46A system and -1.04 kcal/mol for the E46Q system. The resultant free energies are much smaller than the experimental and calculated value of approximately 3 kcal/mol for the native RNaseT1, which suggests that both mutants have greater relative adenine affinities than native RNaseT1. Especially E46Q is calculated to have a larger affinity for adenine than guanine, as we suggested previously from the calculation on the native RNaseT1. Thus, the molecular dynamics/free energy perturbation method may be helpful in protein engineering, directed toward increasing or changing the substrate specificity of enzymes.
authors.affiliation: Department of Pharmaceutical Chemistry, University of California, San Francisco 94143-0446.
authors.forename: S
authors.initials: S
authors.lastname: Hirono
authors.forename: P A
authors.initials: PA
authors.lastname: Kollman
mesh_terms: D000249:Adenosine Monophosphate / Q000494:pharmacology*; D001665:Binding Sites; D004735:Energy Transfer; D006157:Guanosine Monophosphate / Q000494:pharmacology*; D006860:Hydrogen Bonding; D008433:Mathematics; D008958:Models, Molecular; D008968:Molecular Conformation; D016297:Mutagenesis, Site-Directed; D006163:Ribonuclease T1 / Q000037:antagonists & inhibitors* / Q000235:genetics; D013379:Substrate Specificity; D013816:Thermodynamics; D014961:X-Ray Diffraction
chemical_list: C026693:2'-guanylic acid; D000249:Adenosine Monophosphate; D006157:Guanosine Monophosphate; D006163:Ribonuclease T1