SUBSTRATE-BINDING AND CATALYTIC ROLES OF LYS194 IN THE C-TERMINUS IN HUMAN ADENYLATE KINASE BY SITE-DIRECTED RANDOM MUTAGENESIS

Site-directed random mutagenesis of Lys194 residue in the C-terminus o f human adenylate kinase (AK) was performed, and six mutants were anal yzed by steady-state kinetics. K194-mutants variously affected the app arent Michaelis constants (K-m values) for ATP and AMP, although the k (cat) values strikingly decreased. The Lys194 residue appears to inter act not only with MgATP(2-) but also with the AMP(2-) substrates by sa lt bridge formation with a nucleotide and to play a functional role in stabilizing the phosphate-transfer during catalysis. Lys194 could be essential for substrate-holdings and in catalysis and not replaceable to the other amino acids.