STUDIES ON THE REGULATION OF ISOLEUCINE AND VALINE BIOSYNTHESIS IN THE CYANOBACTERIUM SYNECHOCOCCUS LEOPOLIENSIS
Abstract
The regulation of isoleucine and valine biosynthesis in the cyanobacterium S. leopoliensis was studied. Analysis of carbon flow in the organism demonstrated that the first step in the biosynthesis of isoleucine in this organism was the deamination of L-threonine. The enzyme was also shown to be regulated by feedback inhibition by isoleucine. The single acetohydroxyacid synthase of the cyanobacterium with an optimal pH of 7.8 was characterized. The enzyme was shown to be feedback regulated by the end products isoleucine and valine. Leucine was shown to reverse the negative effect of isoleucine but not that of valine. Southern hybridization analysis of the cyanobacterial genome showed the organism to possess a single acetohydroxyacid synthase gene with significant homology to the ilvG gene of E. coli. No hybridization was observed with probes derived from either the ilvBN or the ilvIH genes. A kinetic analysis of the cyanobacterial acetohydroxyacid synthase was conducted on enzyme derived from crude extracts. The enzyme was shown to have a K(,m) of 6.43 x 10('-3) M and a V(,max) of 1.38. A gene was cloned from the cyanobacterial chromosome that specified a protein fully able to complement the functions of the ilvE gene product, transaminase B. The nucleotide sequence of the cyanobacterial gene was determined and compared to the sequences of the ilvE, aspC, and tyrB genes. Significant sequence similarities, at both the nucleotide and amino acid sequence levels, were observed only to the ilvE gene.
Degree
Ph.D.
Subject Area
Molecular biology
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