In Saccharomyces cerevisiae, aspartate kinase (the HOM3 product) regulates the metabolic flux through the threonine biosynthetic pathway through feedback inhibition by the end product. In order to obtain a strain able to produce threonine in a controlled way, we have isolated a mutant allele (HOM3-ts31d) that gives rise to a deregulated aspartate kinase. This allele has been isolated as an extragenic suppressor of ilv1, which confers an Ilv+ phenotype at 37 degrees C but not at 22 degrees C. We have stated that at high temperature the mutant aspartate kinase is slightly more deregulated and shows a higher specific activity, inducing threonine accumulation. The HOM3-ts31d allele carries a mutation that leads to a Ser399 --> Phe substitution in the postulated regulatory region of the enzyme. We have detected other changes in the nucleotide sequence but they are also present in the parental strain, reflecting the genetic differences between different wild-type strains. A sequence comparison among all the reported mutant aspartate kinases suggests that not all residues involved in regulation of the activity are clustered in the so-called regulatory domain, as is the case of that mutated in AK-R7, another deregulated aspartate kinase obtained with the same strategy of ilv1 suppression.

• PMID: 15645479
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Journal Article | Research Support, Non-U.S. Gov't

Authors

Velasco I, Arévalo-Rodríguez M, Marina P, Calderón IL

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