Bacillus methanolicus pyruvate carboxylase and homoserine dehydrogenase I and II and their roles for L-lysine production from methanol at 50 degrees C

Appl Microbiol Biotechnol. 2010 Jul;87(3):951-64. doi: 10.1007/s00253-010-2559-6. Epub 2010 Apr 7.

Abstract

We here present the pyc gene encoding pyruvate carboxylase (PC), and the hom-1 and hom-2 genes encoding two active homoserine dehydrogenase (HD) proteins, in methylotrophic Bacillus methanolicus MGA3. In general, both PC and HD are regarded as key targets for improving bacterial L-lysine production; PC plays a role in precursor oxaloacetate (OAA) supply while HD controls an important branch point in the L-lysine biosynthetic pathway. The hom-1 and hom-2 genes were strongly repressed by L-threonine and L-methionine, respectively. Wild-type MGA3 cells secreted 0.4 g/l L-lysine and 59 g/l L-glutamate under optimised fed batch methanol fermentation. The hom-1 mutant M168-20 constructed herein secreted 11 g/l L-lysine and 69 g/l of L-glutamate, while a sixfold higher L-lysine overproduction (65 g/l) of the previously constructed classical B. methanolicus mutant NOA2#13A52-8A66 was accompanied with reduced L-glutamate production (28 g/l) and threefold elevated pyc transcription level. Overproduction of PC and its mutant enzyme P455S in M168-20 had no positive effect on the volumetric L-lysine yield and the L-lysine yield on methanol, and caused significantly reduced volumetric L-glutamate yield and L: -glutamate yield on methanol. Our results demonstrated that hom-1 represents one key target for achieving L-lysine overproduction, PC activity plays an important role in controlling L-glutamate production from methanol, and that OAA precursor supply is not a major bottleneck for L-lysine overproduction by B. methanolicus.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Bacillus / enzymology*
  • Bacillus / genetics
  • Bacillus / metabolism
  • Bacterial Proteins / genetics
  • Bacterial Proteins / metabolism*
  • Cloning, Molecular
  • Fermentation
  • Glutamic Acid / metabolism
  • Homoserine Dehydrogenase / genetics
  • Homoserine Dehydrogenase / metabolism*
  • Hot Temperature
  • Lysine / biosynthesis*
  • Methanol / metabolism*
  • Methionine / metabolism
  • Molecular Sequence Data
  • Mutation
  • Pyruvate Carboxylase / genetics
  • Pyruvate Carboxylase / metabolism*
  • Threonine / metabolism

Substances

  • Bacterial Proteins
  • Threonine
  • Glutamic Acid
  • Methionine
  • Homoserine Dehydrogenase
  • Pyruvate Carboxylase
  • Lysine
  • Methanol

Associated data

  • GENBANK/DQ025534
  • GENBANK/GU377284
  • GENBANK/GU377285