Functional and evolutionary relationship between arginine biosynthesis and prokaryotic lysine biosynthesis through alpha-aminoadipate.
Our previous studies revealed that lysine is synthesized through alpha-aminoadipate in an extremely thermophilic bacterium, Thermus thermophilus HB27. Sequence analysis of a gene cluster involved in the lysine biosynthesis of this microorganism suggested that the conversion from alpha-aminoadipate to lysine proceeds in a way similar to that of arginine biosynthesis. In the present study, we cloned an argD homolog of T. thermophilus HB27 which was not included in the previously cloned lysine biosynthetic gene cluster and determined the nucleotide sequence. A knockout of the argD-like gene, now termed lysJ, in T. thermophilus HB27 showed that this gene is essential for lysine biosynthesis in this bacterium. The lysJ gene was cloned into a plasmid and overexpressed in Escherichia coli, and the LysJ protein was purified to homogeneity. When the catalytic activity of LysJ was analyzed in a reverse reaction in the putative pathway, LysJ was found to transfer the epsilon-amino group of N(2)-acetyllysine, a putative intermediate in lysine biosynthesis, to 2-oxoglutarate. When N(2)-acetylornithine, a substrate for arginine biosynthesis, was used as the substrate for the reaction, LysJ transferred the delta-amino group of N(2)-acetylornithine to 2-oxoglutarate 16 times more efficiently than when N(2)-acetyllysine was the amino donor. All these results suggest that lysine biosynthesis in T. thermophilus HB27 is functionally and evolutionarily related to arginine biosynthesis.
Miyazaki
J
J
Biotechnology Research Center, The University of Tokyo, 1-1-1 Yayoi, Bunkyo-ku, Tokyo 113-8657, Japan.
Kobashi
N
N
Nishiyama
M
M
Yamane
H
H
D015074:2-Aminoadipic Acid / Q000378:metabolism*; D001120:Arginine / Q000096:biosynthesis*; D001426:Bacterial Proteins; D003001:Cloning, Molecular; D004269:DNA, Bacterial / Q000737:chemistry; D004591:Electrophoresis, Polyacrylamide Gel; D004926:Escherichia coli / Q000235:genetics; D019143:Evolution, Molecular; D007656:Ketoglutaric Acids / Q000378:metabolism; D007700:Kinetics; D008239:Lysine / Q000031:analogs & derivatives / Q000096:biosynthesis* / Q000378:metabolism; D009952:Ornithine / Q000378:metabolism; D017422:Sequence Analysis, DNA; D016963:Thermus thermophilus / Q000378:metabolism; D000637:Transaminases / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism
D001426:Bacterial Proteins; D004269:DNA, Bacterial; D007656:Ketoglutaric Acids; C002315:N(alpha)-acetyllysine; D015074:2-Aminoadipic Acid; D001120:Arginine; D009952:Ornithine; C434051:LysJ protein, Thermus thermophilus; D000637:Transaminases; D008239:Lysine