L-Lysine biosynthetic pathway of Methylophilus methylotrophus and construction of an L-lysine producer.
Previously, we showed that the enzymes aspartokinase (AK) and dihydrodipicolinate synthase (DDPS), which are involved in L-lysine biosynthesis in the Gram-negative obligate methylotroph Methylophilus methylotrophus AS1, were inhibited by allosteric effectors, including L-lysine. To elucidate further the regulation of L-lysine biosynthesis in M. methylotrophus, we cloned the genes encoding three other enzymes involved in this pathway, L-aspartate-beta-semialdehyde dehydrogenase, dihydrodipicolinate reductase (DDPR) and diaminopimelate decarboxylase, and examined their properties. DDPR was markedly inhibited by L-lysine. Based on this and our previous results, we constructed an L-lysine-producing strain of M. methylotrophus by introducing well-characterized genes encoding desensitized forms of AK and DDPS, as well as dapB (encoding DDPR) from Escherichia coli, using a broad host range plasmid. L-Lysine production was significantly increased by employing an S-(2-aminoethyl)-L-cysteine (L-lysine analog)-resistant mutant as the host. This derivative accumulated L-lysine at a concentration of 1 g l(-1) of medium using methanol as a carbon source.
Tsujimoto
Nobuharu
N
Fermentation and Biotechnology Laboratories, Ajinomoto Co., Inc., Kawasaki-ku, Japan.
Gunji
Yoshiya
Y
Ogawa-Miyata
Yuri
Y
Shimaoka
Megumi
M
Yasueda
Hisashi
H
D001223:Aspartate-Semialdehyde Dehydrogenase / Q000235:genetics*; D002262:Carboxy-Lyases / Q000235:genetics*; D050746:Dihydrodipicolinate Reductase / Q000235:genetics*; D015971:Gene Expression Regulation, Enzymologic / Q000235:genetics; D005822:Genetic Vectors / Q000235:genetics; D008239:Lysine / Q000096:biosynthesis*; D000432:Methanol / Q000378:metabolism; D020630:Methylophilus methylotrophus / Q000201:enzymology*; D010957:Plasmids / Q000096:biosynthesis; D015202:Protein Engineering / Q000379:methods
D050746:Dihydrodipicolinate Reductase; D001223:Aspartate-Semialdehyde Dehydrogenase; D002262:Carboxy-Lyases; C020031:diaminopimelic acid decarboxylase; D008239:Lysine; D000432:Methanol