Studies on carboxylases. X.- Enzymatic inhibition of pyruvic carboxylase by the antibiotics of protein nature

Abstract

Continuing the study of the enzymatic inhibition of pyruvic carboxylase by antibiotics of therapeulic importance, the antibiotics bacitracin, polymixine B, tyrotrycin, viomycin and chloramphenicol have been tested. The results obtained have been the following:

1) Bacilracin. Inhibits considerable and substrate concentration has a marked influence on the quantity of inhibition. Theoretical consideration suggests a reversible inhibition of a competitive kind and the possibility that the antibiotic unites with the enzyme in positions which impede the access of the substrate lo the active centre of the enzymatic protein; operating with bacitracin submitted lo gentle hydrolysis, in conditions in which it is know to liberate sulphhydryl groups, a marked increase in inhibitory capacity has been noted. Previous incubation of the antibiotic with the enzyme causes an increased inhibition.

2) Polymyxine B. Appreciably inhibits pyruvic carboxylase and inhibition is reversible with respect to the substrate. The mechanism of inhibition does not seem to fit into any of the types established in the theoretical treatise of Lineweaver and Burke for enzymatic inhibition. The results suggest the formation of a complex, and that a certain state of saturation of the enzyme with the antibiotic is reached, in such a manner that an increase in concentration of the latter does not show an increase in inhibition.

3) Tyrotricin. It inhibits strongly and the inhibition is markedly influenced by the concentration of substrate. Theoretically a reversible but not competitive mechanism with respect to the substrate is indicated. The addition of excess of coenzyme has no effect on inhibition.

4) Viomycin. Inhibits by a coupling mechanism that is, it unites with the free enzyme.

5) Chloramphenicol. Inhibition by this antibiotic is irreversible with respect to the substrate, on the other hand, addition of cocarboxylase markedly reduces inhibition, addition of thiamine has not the same effect.