Cysteine preactivation

The CSA forming enzyme activity was not detected in the preparations after acetone precipitation without preactivation and the inactive enzyme was activated by the anaerobic incubation in the presence of L-cysteine. As shown in Fig. lA and B, the activation was dependent on the substrate concentration and the incubation temperature. The maximum activation was observed when the enzyme was preincubated with 10 mM L-cysteine at 37 for 30 min. Simillar anaerobic activation of the enzyme by L-cysteine was observed either in the presence of or in the absence of protein-A, suggesting that protein-A did not associate with the enzyme activation. 

The activation effect of analogues of L-cysteine such as S-methyl-L-cysteine, DL-homocysteine, cysteamine and N-acetyl-L-cysteine were examined. Since some compounds among of them exhibited a inhibitory effect on the enzyme activity, the preactivation of enzyme was carried out in 0.05 ml of reaction mixture, and then the mixture was diluted to 1 ml with prewarmed buffer solution containing 1 mM L-[35s]cysteine and protein-A, followed by the aerobic incubation under standard conditions. Under these conditions no significant inhibition by the additions was observed. The enzyme was activated by all analogues of L-cysteine. Carboxymethy1-L-cysteine and carboxyethyl-L-cysteine were most effective (13). 

---