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 g l−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.