scholarly journals Enhanced production of gamma-aminobutyrate (GABA) in recombinant Corynebacterium glutamicum by expressing glutamate decarboxylase active in expanded pH range

2015 ◽  
Vol 14 (1) ◽  
Author(s):  
Jae Woong Choi ◽  
Sung Sun Yim ◽  
Seung Hwan Lee ◽  
Taek Jin Kang ◽  
Si Jae Park ◽  
...  
Keyword(s):  
Corynebacterium Glutamicum ◽  
Glutamate Decarboxylase ◽  
Enhanced Production ◽  
Ph Range

Disruption of genes for the enhanced biosynthesis of α-ketoglutarate in Corynebacterium glutamicum

2012 ◽  
Vol 58 (3) ◽  
pp. 278-286 ◽  
Author(s):  
Jae-Hyung Jo ◽  
Hye-Young Seol ◽  
Yun-Bom Lee ◽  
Min-Hong Kim ◽  
Hyung-Hwan Hyun ◽  
...  
Keyword(s):  
Corynebacterium Glutamicum ◽  
Biosynthetic Pathway ◽  
Isocitrate Lyase ◽  
Glutamate Synthase ◽  
Flask Culture ◽  
Control Strain ◽  
Enhanced Production ◽  
Key Enzyme ◽  
Microbial Strains ◽  
Glyoxylate Pathway

The development of microbial strains for the enhanced production of α-ketoglutarate (α-KG) was investigated using a strain of Corynebacterium glutamicum that overproduces of l-glutamate, by disrupting three genes involved in the α-KG biosynthetic pathway. The pathways competing with the biosynthesis of α-KG were blocked by knocking out aceA (encoding isocitrate lyase, ICL), gdh (encoding glutamate dehydrogenase, l-gluDH), and gltB (encoding glutamate synthase or glutamate-2-oxoglutarate aminotransferase, GOGAT). The strain with aceA, gltB, and gdh disrupted showed reduced ICL activity and no GOGAT and l-gluDH activities, resulting in up to 16-fold more α-KG production than the control strain in flask culture. These results suggest that l-gluDH is the key enzyme in the conversion of α-KG to l-glutamate; therefore, prevention of this step could promote α-KG accumulation. The inactivation of ICL leads the carbon flow to α-KG by blocking the glyoxylate pathway. However, the disruption of gltB did not affect the biosynthesis of α-KG. Our results can be applied in the industrial production of α-KG by using C. glutamicum as producer.

scholarly journals Phagocytosis by Human Monocytes

Blood ◽  
1968 ◽  
Vol 32 (3) ◽  
pp. 423-435 ◽  
Author(s):  
MARTIN J. CLINE ◽  
ROBERT I. LEHRER
Keyword(s):  
Rna Synthesis ◽  
Glucose Utilization ◽  
Human Monocytes ◽  
Igg Antibody ◽  
Enhanced Production ◽  
Serum Fractions ◽  
Particle Ingestion ◽  
Ph Range ◽  
Total Glucose

Abstract Human monocytes isolated from peripheral blood phagocytized polystyrene particles, Escherichia coli, Staphylococcus aureus, Candida albicans, Cryptococcus neoformans, and red cells coated with IgG antibody in vitro. The ingestion of fungi and bacteria required serum or serum fractions. Ingestion of antibody-coated erythrocytes was inhibited by whole serum or IgG, but not by IgM or albumin. Adherence to a surface appeared to be critical for erythrophagocytosis but not for ingestion of bacteria. Phagocytosis occurred over a broad pH range and in the presence of inhibitors of oxidative metabolism and of RNA synthesis. Phagocytosis was significantly depressed by inhibitors of glycolysis (iodoacetate and fluoride) and by chloroquine at 1 x 10-5M. Particle ingestion by monocytes was accompanied by enhanced production of C14O2 from glucose-1-C14, but was not associated with a detectable increase in total glucose utilization, incorporation of radioactive uridine into RNA, or incorporation of amino acids into protein.

Expanding the active pH range of Escherichia coli glutamate decarboxylase by breaking the cooperativeness

2013 ◽  
Vol 115 (2) ◽  
pp. 154-158 ◽  
Author(s):  
Ngoc Anh Thu Ho ◽  
Chen Yuan Hou ◽  
Woo Hyun Kim ◽  
Taek Jin Kang
Keyword(s):  
Escherichia Coli ◽  
Glutamate Decarboxylase ◽  
Ph Range

scholarly journals Functional Caracterization of CapBCA in Controlling Poly-γ-Glutamic Acid Synthesis in Corynebacterium Glutamicum

2021 ◽  
Author(s):  
Guoqiang Xu ◽  
Jiyue Wang ◽  
Luning Gu ◽  
Yaxin Zhu ◽  
Jian Zha ◽  
...  
Keyword(s):  
Cell Membrane ◽  
Glutamic Acid ◽  
Environmental Protection ◽  
Corynebacterium Glutamicum ◽  
Bioinformatics Analysis ◽  
Acid Synthesis ◽  
Transcription Level ◽  
Enhanced Production ◽  
Medium Level ◽  
Shed Light

Abstract Background Poly-γ-glutamic acid (γ-PGA) is a natural anionic biopolymer widely used in various fields, including medicine, food, cosmetics, and environmental protection. The γ-PGA synthase complex, CapBCA, is the only polyprotein complex responsible for γ-PGA synthesis. However, systematic and in-depth research on the function of each component involved in γ-PGA synthesis is scarce, which limits enhanced production of γ-PGA. Results To address this limitation, γ-PGA synthase components were localized, and their functions associated with γ-PGA synthesis were investigated in Corynebacterium glutamicum. Bioinformatics analysis and confocal microscopic observations of CapB-sfGFP, CapC-sfGFP, and CapA-sfGFP proteins revealed that γ-PGA synthase components CapB, CapC, and CapA were all localized on the cell membrane. More importantly, γ-PGA was detected only when CapB, CapC, and CapA were expressed in combination in C. glutamicum. Furthermore, enhancement of CapB or CapC transcription levels (from low to high) and maintaining medium-level CapA transcription led to 35.44% and 76.53% increase in γ-PGA yield (γ-PGA yield-to-biomass), respectively. However, maintaining medium-level CapB and CapC transcription, and moderate enhancement of CapA transcription level (from low to medium) led to 35.01% increase in γ-PGA yield, whereas a further increase in CapA expression (from medium to high) led to 10.36% decrease in γ-PGA yield. Notably, CapC had the greatest influence (accounting for 68.24%) on γ-PGA synthesis. Conclusions The present study determined the membrane localization of γ-PGA synthase components, CapB, CapC, and CapA, in C. glutamicum and confirmed the significance of these components in γ-PGA production. Furthermore, CapC was found to have the greatest influence on controlling γ-PGA synthesis. These findings shed light into the effect of γ-PGA synthase component expression on γ-PGA synthesis, and provide insights for further improvement in γ-PGA production.

Characterization and Mutagenesis of a Novel Mycobacterium Smegmatis -Derived Glutamate Decarboxylase Active at Neutral pH

2021 ◽  
Author(s):  
Yudi Li ◽  
Guiying Chen ◽  
Fanglan Ge ◽  
Ting Dang ◽  
Yao Ren ◽  
...  
Keyword(s):  
Enzymatic Activity ◽  
Mycobacterium Smegmatis ◽  
Glutamate Decarboxylase ◽  
Aminobutyric Acid ◽  
High Catalytic Activity ◽  
Wild Type ◽  
Rapid Loss ◽  
Neutral Ph ◽  
Key Enzyme ◽  
Ph Range

Abstract γ-aminobutyric acid (GABA) has various physiological functions and is widely used in medicine, food, and other fields. Glutamate decarboxylase (GAD) is a key enzyme that catalyzes the decarboxylation of L-glutamate to synthesize GABA. However, the industrial application of microorganism-derived GAD is limited by its rapid loss of enzymatic activity with pH approaching neutrality. In this study, a novel glutamate decarboxylase, GAD MSM, from Mycobacterium smegmatis was overexpressed and purified. On the basis of homologous modeling and substrate molecular docking, several GAD MSM mutants were constructed, and their enzymatic properties were analyzed. The results showed that the optimal pH of wild-type GAD MSM is 5.4; at pH 6.2, 22.8% enzymatic activity was retained; the T211 amino acid residue and C-terminal deletion mutant GAD MSMΔC showed relatively high catalytic activity in a pH range of 5.0–7.0. The V max and K m values of GAD MSMΔC were 14.69 and 5.70, respectively, at pH 5.5, and 9.87 and 6.17, respectively, at pH 7.0. Compared with the wild-type GAD, GAD MSMΔC still maintained higher affinity and enzymatic activity of the substrate, maintaining 78.5% of the highest enzymatic activity even at pH 7.0, which is the highest reported activity retention for GAD under neutral pH condition. Therefore, GAD MSMΔC can be used for the transformation of high-yielding strains and industrial production of GABA.

Sign in / Sign up

Export Citation Format

Share Document