Effects of the amplification of the genes coding for the L-threonine biosynthetic enzymes on the L-threonine production from methanol by a gram-negative obligate methylotroph, Methylobacillus glycogenes.
We constructed recombinant plasmids carrying the genes coding for the L-threonine biosynthetic enzymes, the hom gene, the hom-thrC genes, and the thrB genes, of a gram-negative obligate methylotroph, Methylobacillus glycogenes, and examined the effects of them on the production of L-threonine from methanol. The hom gene, which encodes the homoserine dehydrogenase, and the hom-thrC genes, containing the gene coding for threonine synthase together with the hom gene, were cloned from a wild-type strain, and the thrB gene encoding the desensitized homoserine kinase was cloned from an L-threonine-producing mutant, ATR80. The recombinant plasmids were transferred into ATR80 and its L-isoleucine auxotroph, A513, by conjugation. Amplification of the genes coding for the L-threonine biosynthetic enzymes elevated the activities of the L-threonine biosynthetic enzymes of the transconjugants 10- to 30-fold over those of the strains containing only vectors. The L-threonine production from methanol in test-tube cultivation was increased about 30% and 40% by the amplification of the hom gene and the hom-thrC gene respectively, and it was slightly increased by that of the thrB gene. The effects of gene amplification were confirmed by the cultivation in 5-1 jar fermentors. The best producer, an A513 transconjugant containing the plasmid carrying the hom-thrC genes, produced 16.3 g/l L-threonine for 72 h.
Motoyama
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H
Tokyo Research Laboratory, Kyowa Hakko Kogyo Co. Ltd., Japan.
Yano
H
H
Ishino
S
S
Anazawa
H
H
Teshiba
S
S
D001709:Biotechnology; D019757:Carbon-Oxygen Lyases; D002874:Chromosome Mapping; D003001:Cloning, Molecular; D005784:Gene Amplification; D005798:Genes, Bacterial; D006715:Homoserine Dehydrogenase / Q000235:genetics; D008190:Lyases / Q000235:genetics; D000432:Methanol / Q000378:metabolism; D008772:Methylococcaceae / Q000235:genetics* / Q000378:metabolism*; D010957:Plasmids / Q000235:genetics; D013912:Threonine / Q000096:biosynthesis*; D014170:Transformation, Genetic
D013912:Threonine; D006715:Homoserine Dehydrogenase; D008190:Lyases; D019757:Carbon-Oxygen Lyases; C023160:threonine synthase; D000432:Methanol