A strain (MIAR33) of Saccharomyces cerevisiae containing a threonine deaminase [L-threonine hydrolyase (deaminating) EC 4.2.1.161 with decreased feedback sensitivity has been shown to have a specific activity of acetohydroxy acid synthetase higher than that of the parent strain (MDIll) when both are grown on minimal medium. When strain MAR33 is grown on minimal medium supplemented only with isoleucine, the specific activity of the synthetase is reduced to that found in the parent strain. Another strain, D106-lA, contains a nonsense mutation in the middle of the gene for threonine deaminase. When this strain is grown on minimal medium containing appropriate supplements (which include a nonrepressing concentration of isoleucine), or on minimal medium supplemented with isoleucylglycine (which acts as a limiting source of isoleucine), acetohydroxy acid synthetase remains repressed. Leucine limitation causes partial derepression. With the reversion of the nonsense mutation, either intragenically or via a suppressor for the mutation, partial derepression of the synthetase returns. When D106-lA is diploidized with either M15, a mutant lacking the synthetase, or MD9, a strain containing the enzyme, normal, partially derepressed, values for this enzyme are found. This indicates that threonine deaminase is necessary for derepression, and that it possibly acts as an inducer.The involvement of a biosynthetic enzyme of a specific aminoacid pathway in repression of other enzymes of the same pathway has been considered by several investigators (1-7).Our possession of several mutants affected in the allosteric properties of threonine deaminase permitted us to explore the involvement of this enzyme in the repression of acetohydroxy acid synthetase in Saccharomyces cerevisiae. Threonine deaminase and acetohydroxy acid synthetase are the first two enzymes in the isoleucine-valine biosynthetic pathway. The properties of these two enzymes in S. cerevisiae have been examined (8-10). Regulation of the isoleucinevaline pathway has been examined in S. cerevisiae (11, 12), in Escherichia coli (13-15), and in Salmonella typhimurium (16). All of these organisms show multivalent repression of at least some of the enzymes of the pathway. Full repression occurs only in the presence of all three aminio acids: isoleucine, valine, and leucine. Starvation for one of these compounds causes derepression of the enzymes of the pathway, even in the presence of an excess of the other two ( 1).
METHODS AND RESULTSThtreonine deaminase mutated in its allosteric properties and the repression of acetohydroxy acid synthetase MAR33, a haploid strain of S. cerevisiae, contains a single mutation that renders threonine deaminiase 100-fold less sensitive to isoleucine inhibition than the enzyme of MD11 (the parent strain) (17). Parent and mutant strains were grown inl minimal and repressing medium, which is minimal medium supplemented with isoleucine, valine, and leucine, each at 5 mM\. The cultures were made permeable with toluene, and the acetohydroxy a...