The proteolytic system of the yeast Kluyveromyces lactis.

Major proteolytic activities were characterized in the yeast K. lactis NRRL 1118, grown in chemostat cultures. This yeast expressed proteolytic activities similar to those found in S. cerevisiae. This fact was particularly evident in the case of proteases such as PrA, PrB and CpY with regard to substrate specificity, activation at pH 5. 0 and inhibition patterns. The presence of a CpS activity could not be detected in either fresh or activated cell-free extracts by using the dipeptide N-Cbz-Gly-Leu, even in the presence of Zn(+2). On the other hand, K. lactis exhibits at least two major intracellular Ap activities different from those reported in other yeasts, and these seem to be carried out by closely related proteins. These activities corresponded to molecular masses of about 60 kDa, close pI values, and a similar behaviour in non-denaturing polyacrylamide electrophoresis. Both activities were enhanced by Co(+2) and inhibited by EDTA. Among different aminoacyl-p-NAs, they preferentially hydrolysed Lys-p-NA. No increase of Ap activity was obtained by incubation of extracts at acid pH. The maximum PrA and PrB activities detected in N-limited cultures were six-fold higher than those expressed under C- or P-limitation. The effect of culture conditions on the Cp and Ap expression was much less pronounced in comparison with PrA and PrB activities, Ap levels even being slightly higher in C-limited cells. This fact suggests that hydrolysis of protein to peptides might be the limiting step in the pathway of general protein degradation in the vacuole.