Studies on carboxylases. VIII.- Enzymatic inhibition of pyruvic carboxylase by antibiotics derived from the naphthotacenic nucleus

Abstract

A study has been made of the inhibitory action of the antibiotics tetracycline, chlorotetracycline and oxytetracycline on purified pyruvic carboxylase. The velocity of decarboxylation of pyruvic acid by the enzyme preparation at different antibiotic concentrations was determined manometrically.

The graphs which represent v/v, as regards the molar concentration of antibiotic, present two very marked points of inflection in the three derivatives of the naphthacenic nucleus, wich indicates that these three compouds behave in a similar way qualitatively and quantitatively.

The substrate has in all cases a protective action on the enzyme activity; in the case of tetracycline and of chlorotetracycline, a ten-fold increase in substrate concentration reduces the inhibition to approximately one half, while in the case of oxytetracycline the protective action of the substrate is less marked.

The addition of inert proteins considerably reduces the inhibition and thus employing crude preparations of the enzyme a much smaller inhibition is reached than with the purified enzyme : this indicates that the three antibiotics mentioned unite with proteins to form non-specific complexes, complexes which may facilitate distribution in the organism through the plasmatic proteins.

The divalent cations magnesium and manganese have a marked effect on the inhibition: the addition of 5 x 10-4 mols by liter of ions reduces the inhibition by half; on the other hand in the least. These results suggest that the mechanism of inhibition is complex, due to the funcional groups common to the three antibiotics studied; part of the inhibition is caused by non-specific interaction with the protein part of the enzyme and another part is due to union with the divalent ions which the enzyme requires for its functioning. Results with cysteine indicate that there is no block nor oxidation of the sulp-hydryl groups of the enzyme.