Glycinamide Ribonucleotide Transformylase (GART) is an important enzyme for the biosynthesis of purine bases for DNA. It is also an attractive candidate for targeting with antineoplastic or antimicrobial agents. Understanding of structure-function relations in GART is a first step for rational drug design. The availability of high resolution crystal structure data and detailed kinetic mechanisms describing the enzymatic reaction make GART an ideal model system to investigate significant domain-domain interactions that are important for structure formation and catalysis. We propose to theoretically calculate the pKa values of all ionizable groups of GART using a novel computational approach which is based on solving the linearized Poisson-Boltzmann equation. We expect to delineate the role of the ionizable groups that are responsible for the monomer-dimer transition and to understand the role His54, His7, Glu70, Tyr67, Tyr78, Tyr115 and other titratable residues in the mechanism of the transition. Since this calculation will utilize the X-ray structures of GART we expect to elucidate the significance of the conformational changes upon dimerization. We propose to apply a combination of Monte Carlo and Stochastic dynamics methods (MC/SD) to model the pH-dependent disordered loop-ordered loop/helix transition in residues 110-131 of the Glu70Ala GART mutant and the effect of the local conformational transition in the protein overall and its catalytic activity.
