Enhancement of L-lysine production in methylotroph Methylophilus methylotrophus by introducing a mutant LysE exporter.
The obligate methylotroph Methylophilus methylotrophus AS1 expressing a mutant form of dapA (dapA24) encoding a dihydrodipicolinate synthase desensitized from feedback inhibition by L-lysine could secrete L-lysine into the medium, but also maintained a high concentration of intracellular L-lysine. To improve the yield from excretion, we attempted to introduce an L-lysine/L-arginine exporter (LysE) from Corynebacterium glutamicum 2256 into M. methylotrophus. We were unable to stably transform M. methylotrophus with a plasmid expressing the wild type lysE gene, but happened to obtain a transformant carrying a spontaneously mutated lysE gene (designated lysE24) which could induce L-lysine production even in the wild type strain. The transformant also possessed increased tolerance to S-(2-aminoethyl)-L-cysteine (an L-lysine analog). lysE24 has a single-base insertion mutation in the middle of the lysE gene, and its product is presumably quite different in structure from wild-type LysE. When lysE24 was introduced into an L-lysine producer of M. methylotrophus carrying dapA24, the level of intracellular L-lysine fell. During fermentation, M. methylotrophus carrying both lysE24 and dapA24 produced 10-fold more L-lysine (11.3 gl(-1) in jar-fermentation) than the parent producer carrying only dapA24 or lysE24. These results show the importance of the factor (lysE24) involved in the excretion of L-lysine on its overproduction in M. methylotrophus.
Gunji
Yoshiya
Y
Fermentation and Biotechnology Laboratories, Ajinomoto Co., Inc. Kawasaki, Japan.
Yasueda
Hisashi
H
D026923:Amino Acid Transport Systems, Basic / Q000235:genetics; D001426:Bacterial Proteins / Q000235:genetics; D019149:Bioreactors; D008239:Lysine / Q000235:genetics / Q000378:metabolism*; D000432:Methanol / Q000378:metabolism; D020630:Methylophilus methylotrophus / Q000235:genetics / Q000378:metabolism*; D008969:Molecular Sequence Data; D009154:Mutation; D014169:Transformation, Bacterial / Q000235:genetics* / Q000502:physiology
D026923:Amino Acid Transport Systems, Basic; D001426:Bacterial Proteins; C104616:LysE protein, Corynebacterium glutamicum; D008239:Lysine; D000432:Methanol