INITIAL-STATE OF AN ENZYMATIC-REACTION - THEORETICAL PREDICTION OF COMPLEX-FORMATION IN THE ACTIVE-SITE OF RNASE T1

A computer model for the hydrated complex between the enzyme ribonucle ase (RNase) T1 and its substrate guanylyl-3',5'-guanosine has been ref ined using molecular dynamics simulation and quantum chemical calculat ions. Actual protonation states of the most important residues at the active site in the presence of the substrate were derived from publish ed NMR titrations and pH-dependent kinetic studies, which were confirm ed by independent Monte Carlo calculations (manuscript in preparation) . The molecular dynamics trajectory has been analyzed to theoretically capture the initial point of the enzymatic reaction pathway. The chan ges in the charge distribution of the most relevant part of the enzyme -substrate complex have been checked by the CNDO/2-spd technique. The initial point of the enzymatic reaction pathway has been found to corr espond as expected, to the strained conformation of the ''substrate active site side chains'' complex. His40, Glu58, Arg77, and His92 whic h are primarily involved in the enzymatic activity show hydrogen bond contacts to the substrate. In this scheme, Glu58 plays the role of gen eral base and His92 acts as the general acid in the reaction pathway, while the other two residues stabilize the initial state of the reacti on electrostatically.