title: Improved production of secreted heterologous enzyme in Bacillus subtilis strain MGB874 via modification of glutamate metabolism and growth conditions.
abstract: BACKGROUND
  The Bacillus subtilis genome-reduced strain MGB874 exhibits enhanced production of exogenous extracellular enzymes under batch fermentation conditions. We predicted that deletion of the gene for RocG, a bi-functional protein that acts as a glutamate dehydrogenase and an indirect repressor of glutamate synthesis, would improve glutamate metabolism, leading to further increased enzyme production. However, deletion of rocG dramatically decreased production of the alkaline cellulase Egl-237 in strain MGB874 (strain 874∆rocG).
  
  
  RESULTS
  Transcriptome analysis and cultivation profiles suggest that this phenomenon is attributable to impaired secretion of alkaline cellulase Egl-237 and nitrogen starvation, caused by decreased external pH and ammonium depletion, respectively. With NH3-pH auxostat fermentation, production of alkaline cellulase Egl-237 in strain 874∆rocG was increased, exceeding that in the wild-type-background strain 168∆rocG. Notably, in strain 874∆rocG, high enzyme productivity was observed throughout cultivation, possibly due to enhancement of metabolic flux from 2-oxoglutarate to glutamate and generation of metabolic energy through activation of the tricarboxylic acid (TCA) cycle. The level of alkaline cellulase Egl-237 obtained corresponded to about 5.5 g l-1, the highest level reported so far.
  
  
  CONCLUSIONS
  We found the highest levels of production of alkaline cellulase Egl-237 with the reduced-genome strain 874∆rocG and using the NH3-pH auxostat. Deletion of the glutamate dehydrogenase gene rocG enhanced enzyme production via a prolonged auxostat fermentation, possibly due to improved glutamate synthesis and enhanced generation of metabolism energy.
authors: Biological Science Laboratories, Kao Corporation, 2606 Akabane, Ichikai, Haga, Tochigi 321-3497, Japan.
authors: Kenji
authors: K
authors: Manabe
authors: Yasushi
authors: Y
authors: Kageyama
authors: Takuya
authors: T
authors: Morimoto
authors: Eri
authors: E
authors: Shimizu
authors: Hiroki
authors: H
authors: Takahashi
authors: Shigehiko
authors: S
authors: Kanaya
authors: Katsutoshi
authors: K
authors: Ara
authors: Katsuya
authors: K
authors: Ozaki
authors: Naotake
authors: N
authors: Ogasawara
mesh_terms: D000641:Ammonia / Q000378:metabolism; D001412:Bacillus subtilis / Q000254:growth & development / Q000378:metabolism*; D044602:Cellulases / Q000235:genetics / Q000378:metabolism*; D002952:Citric Acid Cycle; D015536:Down-Regulation; D020869:Gene Expression Profiling; D005969:Glutamate Dehydrogenase / Q000235:genetics / Q000378:metabolism; D018698:Glutamic Acid / Q000378:metabolism*; D006863:Hydrogen-Ion Concentration; D007656:Ketoglutaric Acids / Q000378:metabolism; D017384:Sequence Deletion
chemical_list: D007656:Ketoglutaric Acids; D018698:Glutamic Acid; D000641:Ammonia; D005969:Glutamate Dehydrogenase; D044602:Cellulases