Molecular Interactions of Uncompetitive Inhibitors with Bovine Liver Glutamate Dehydrogenase: SODIUM ALKYL SULFATES

Abstract

Abstract A series of sodium n-alkyl sulfate esters (6, 8, 10, 12, 14, and 16 carbon atoms) were synthesized from the respective alcohols. The relative potency (negative logarithm of detergent concentration producing 50% inhibition) of these sodium dodecyl sulfate analogs for inhibition of bovine liver glutamate dehydrogenase was a linear function of the number of carbon atoms in the detergent. This linear correlation indicated that hydrophobic bonding interactions were a major force in the combination of inhibitor with enzyme. This was substantiated by the observation that a linear correlation was also obtained between log (1/I50) and log P. P was the partition coefficient for the inhibitor between the phases 1-octanol and water. Lengthening the alkyl chain of the anionic detergents beyond 14 carbon atoms did not bring an additional enhancement of inhibition. This indicated a possible size limitation of about 23 A (extended length of C14 analog) or 13 A (for a folded configuration of the detergent) at the detergent binding site on the enzyme. Inhibition occurred without denaturation since changes in protein fluorescence occurred only at much higher cencentrations than those used in the kinetic studies. Denaturation activity by the detergents (log 1/F50, F was the concentration of detergent producing 50% of the change in protein tryptophanyl fluorescence) was also linearly related to the number of carbon atoms in the sulfate ester. Double reciprocal plots of velocity against l-glutamate concentration indicated that sodium hexyl sulfate was a noncompetitive inhibitor. Sodium octyl sulfate gave mixed kinetics between competitive and uncompetitive inhibition. Sodium decyl, dodecyl, tetradecyl, and hexadecyl sulfates were uncompetitive inhibitors. The inhibition by sodium decyl and dodecyl sulfates involved greater cooperativity than that which occurred with the other analogs. Sulfate ion was observed to be a weak competitive inhibitor (Ki, 0.29 m). Sodium laurate and cetyltrimethylammonium bromide were cooperative uncompetitive inhibitors with respect to l-glutamate. Thus, either a positive or negative charge at one end of the hydrophobic alkyl chain will suffice for uncompetitive inhibition by these detergents.