Asparaginase II of Saccharomyces cerevisiae: Positive selection of two mutations that prevent enzyme synthesis

Abstract

A positive selection method, D-aspartic acid β-hydroxamate resistance, was used to isolate Saccharomyces cerevisiae strains lacking the ability to synthesize asparaginase II. Of 100 such mutant strains, 93 exhibited mutations which were allelic with asp3, a previously characterized mutation. The other seven strains carried a new mutation asp6. The asp6 mutation segregated 2:2 in asp6 x wild-type crosses and assorted from the asp3 mutation in asp6 x asp3 crosses. All seven asp6 mutant isolates reverted at a relatively high frequency, whereas the asp3 mutant isolates did not revert under the same conditions. Various independent asp3 isolates were mated to give heteroallelic diploids, which when sporulated and spread on D-asparagine medium yielded no recombinant strains.