Deletion of genes involved in glutamate metabolism to improve poly-gamma-glutamic acid production in B. amyloliquefaciens LL3

Abstract

Here, we attempted to elevate poly-gamma-glutamic acid (γ-PGA) production by modifying genes involved in glutamate metabolism in Bacillus amyloliquefaciens LL3. Products of rocR, rocG and gudB facilitate the conversion from glutamate to 2-oxoglutarate in Bacillus subtillis. The gene odhA is responsible for the synthesis of a component of the 2-oxoglutarate dehydrogenase complex that catalyzes the oxidative decarboxylation of 2-oxoglutarate to succinyl coenzyme A. In-frame deletions of these four genes were performed. In shake flask experiments the gudB/rocG double mutant presented enhanced production of γ-PGA, a 38 % increase compared with wild type. When fermented in a 5-L fermenter with pH control, the γ-PGA yield of the rocR mutant was increased to 5.83 g/L from 4.55 g/L for shake flask experiments. The gudB/rocG double mutant produced 5.68 g/L γ-PGA compared with that of 4.03 g/L for the wild type, a 40 % increase. Those results indicated the possibility of improving γ-PGA production by modifying glutamate metabolism, and identified potential genetic targets to improve γ-PGA production.

Improved poly-γ-glutamic acid production in Bacillus amyloliquefaciens by modular pathway engineering

“…Commichau and Stülke (2008) pointed out that GS is a trigger enzyme that is active in metabolism and gene expression controlling; thus we hypothesize that the inhibition of glnA gene expression would disrupt the normal metabolic of bacteria thereafter lead to the decrease of γ-PGA production in the NK-anti-glnA and NK-anti-glnA-rocG strains. Zhang et al (2014aZhang et al ( , 2014b have demonstrated that the rocG gene deletion has no effect on γ-PGA production. However, in this study the inhibition of rocG gene expression resulted in the increase of γ-PGA production.…”

“…To carry out Q4 multiple gene deletions or insertions on a single strain, a marker-less gene deletion method was used to construct the gene knockout mutants (Keller et al, 2009;Zhang et al, 2014aZhang et al, , 2014b.…”

Biological Activity and Biosynthesis of Poly-γ-Glutamic Acid by Bacteria of Bacillus Genus

“…Образование γ-ПГК происходит нерибосомально в два этапа: на первом происходит синтез L-и D-глутаминовой кислоты, на втором -их полимеризация в γ-ПГК. Генетическая организация синтетического аппарата B. anthracis и других непатогенных бацилл различна [10]. У B. anthracis синтез γ-ПГК тесно связан с плазмидами pXO1 (180 kb) и pXO2 (95 kb).…”

Metabolic engineering ofBacillus amyloliquefaciensLL3 for enhanced poly‐γ‐glutamic acid synthesis

“…Zhang et al . () deleted the genes rocR and gudB in B. amyloliquefaciens LL3, and the mutant showed 38% increase in the production of γ‐PGA. Feng et al .…”