Summary: The nucleotide sequence of the operon from a proline-hyperproducing mutant of was determined. Two base substitutions were found: one in the structural gene, coding for γ-glutamyl kinase (GK), and a second one in the promoter region of the operon. The former base substitution led to a change of the predicted amino acid at position 117 from an alanine to a valine in GK. This mutation rendered GK 700-fold less sensitive to proline-mediated feedback inhibition than the wild-type enzyme. The other base substitution, a transversion from a G-C to an A-T, was located in the spacer region between the ‘---35’ and ‘---10’ sequences of the promoter, and it increased the transcriptional activity of this operon fourfold. Both these two base substitutions, which were acquired at the step of selecting mutants resistant to a toxic proline analogue, 3,4-dehydroproline, confer upon cells a high proline productivity and an increased osmotolerance.


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