The kinetics of the reversible inhibition of heart lactate dehydrogenase through the formation of the enzyme–oxidized nicotinamide–adenine dinucleotide–pyruvate compound
The inhibition of lactate dehydrogenase at high pyruvate concentration was studied in three ways. First, a rapid decrease in the rate of the enzyme reaction was observed; secondly, the rate of formation of a pyruvate–NAD+ compound was followed by the change in E325; thirdly, the rate of quenching of the protein fluorescence was measured. The data obtained at pH6·0 at different temperatures and ionic strengths as functions of pyruvate, NAD+ and enzyme concentrations show that the extent of inhibition can be correlated with the reversible formation of a compound between pyruvate and enzyme-bound NAD+. It is suggested that the detailed kinetic analysis of the formation of this abortive ternary compound will give pertinent information about properties of the enzyme–NAD+ compound involved in the normal catalytic process.