EFFECTS OF ANOXIC CONDITIONS ON THE ENZYMATIC CONVERSION OF D,L-2-AMINO-THIAZOLINE-4-CARBOXYLIC ACID TO L-CYSTINE

The effects of anoxic conditions on product inhibition and the stabili ty of L-ATC hydrolase were investigated in the conversion of D,L-2-ami no-Delta(2)-thiazoline-4-carboxylic acid (D,L-ATC) to L-cystine using the cell free extract enzyme of Pseudomonas sp. in the presence of hyd roxylamine. At L-cysteine equivalent levels, where one mole of L-cysti ne was counted as two moles of L-cysteine, L-cystine inhibited the L-A TC hydrolase reaction to a greater extent than L-cysteine. In air, the product occurred predominantly as L-cystine (94.9%), whereas in a nit rogen atmosphere the product occurred as a mixture of L-cysteine (39.3 %) and L-cystine (40.7%). As a result, less product inhibition took pl ace in nitrogen. The activity of L-ATC hydrolase was almost fully lost after 20 h of incubation by shaking at 30 degrees C in air, but consi derable activity remained under the anoxic conditions of nitrogen. A k inetic analysis of the reactions confirmed that reduced product inhibi tion and enhanced enzyme stability in nitrogen result in a more effici ent enzyme reaction. The inactivation rate constant (k(1)) was estimat ed to be 0.11 h(-1) in nitrogen and 0.22(-1) in air, indicating that t he stability of L-ATC hydrolase in nitrogen was greater than in air. T he values of the K-p1 and K-p2 constants related to product inhibition were 43.36 mM and 30.48 mM for L-cysteine and 2-cystine, respectively , where higher values were an indication of less product inhibition. T he value of the rate constant (k(2)) for the oxidation of L-cysteine t o L-cystine was 0.09 h(-1) in nitrogen and 1.01 h(-1) in air, suggesti ng that the oxidation of L-cysteine to L-cystine proceeds faster in ai r than in nitrogen.