Evidence for an essential arginine residue at the active site of ATP citrate lyase from rat liver

Publisher Website
Google Scholar
Abstract
Rat liver ATP citrate lyase was inactivated by 2,3-butanedione and phenylglyoxal. Phenylglyoxal caused the most rapid and complete inactivation of enzyme activity in 4-(2-hydroxyethyl)-1-piperazine-ethanesulfonic acid buffer, pH 8. Inactivation by both butanedione and phenylglyoxal was concentration-dependent and followed pseudo-1st-order kinetics. Phenylglyoxal also decreased autophosphorylation (catalytic phosphate) of ATP citrate lyase. Inactivation by phenylglyoxal and butanedione was due to the modification of enzyme arginine residues; the modified enzyme failed to bind to CoA-agarose. The V declined as a function of inactivation, but the Km values were unaltered. The substrates, CoASH and CoASH plus citrate, protected the enzyme significantly against inactivation, but ATP provided little protection. Inactivation with excess reagent modified .apprx. 8 arginine residues/monomer of enzyme. Citrate, CoASH and ATP protected 2-3 arginine residues from modification by phenylglyoxal. Analysis of the data by statistical methods suggested that the inactivation was due to modification of 1 essential arginine residue/monomer of lyase, which was modified 1.5 times more rapidly than were the other arginine residues. Apparently this essential arginine residue is at the CoASH binding site.