Volume 12, Issue 46 (3-2022)
NCMBJ 2022, 12(46): 65-74 |
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Razavipour R, Akhavan Sepahi A, Modarressi M H, Bambai B. Inhibitory role of formate dehydrogenase enzyme in the growth of BL21 industrial bacteria. NCMBJ 2022; 12 (46) :65-74
URL: http://ncmbjpiau.ir/article-1-1456-en.html
URL: http://ncmbjpiau.ir/article-1-1456-en.html
Department Of Biology, Science and Research Branch Islamic Azad University, Tehran, Iran
Abstract: (1734 Views)
Aim and Background: High metabolism occurs in the presence of oxygen with rapid growth rate in a wide range of organisms including bacteria, yeasts or cancer cells. The ability to grow at high yields plays an important role in biotechnology, especially during the production of proteins (preferably recombinant) or metabolic compounds such as organic acids and secondary metabolites. Increasing the growth efficiency of bacteria, especially Escherichia coli, which is the engine of research and industrial activities, is a noble goal in microbial biotechnology. In this study, we sought to find a solution to reduce the amount of CO2 produced and thus increase the production efficiency of cell mass from organic matter in the process of growth and proliferation by examining the metabolic pathways of E. coli.
Materials and Methods: Metabolic pathways documented on the KEGG site were examined with a view to reducing the CO2 production efficiency of formic acid. Two strains of knockout bacteria of K12 origin were obtained from Keio microbial bank. Then, selected strains were cultured in complex (LB) and simple (M9 + Glycerol) media. Cell mass production in different treatments were compared with the standard BL21 strain based on optical absorption at 600 nm.
Results: In the LB complex medium, Escherichia coli mutants (W3866 and W4040) grew faster than BL21 (approximately 5-fold at 8 and 10 h and 3 times at 12 h compared to samples with the dehydrogenase formate gene). However, this difference was more pronounced in the simple M9 medium, as we observed more than 6-fold growth in 24 hours of incubation in mutant samples lacking the FDH gene compared to the BL21 maternal strain.
Discussion: The experiments were completely in line with metabolic predictions and the growth of mutant bacteria was higher than that of BL21. Interestingly, most mutants grew in a simple medium containing glycerol, which showed that glycerol is a very good source for the growth of Escherichia coli bacteria. These results explain the inconsistency of predictions of previous metabolic models that declared glycerol a suitable carbon source for the growth of E. coli, but did not achieve it in practice.
Conclusion: Under normal physiological conditions, E. coli is not able to grow high in glycerol medium. Deletion of formate dehydrogenase gene caused fundamental changes in metabolic process and increased growth rate compared to BL21 strain, which indicates the inhibitory role of this enzyme in increased growth efficiency of E. coli in the presence of glycerol. In addition, the resulting strain can be used to express recombinant proteins with higher efficiency
Materials and Methods: Metabolic pathways documented on the KEGG site were examined with a view to reducing the CO2 production efficiency of formic acid. Two strains of knockout bacteria of K12 origin were obtained from Keio microbial bank. Then, selected strains were cultured in complex (LB) and simple (M9 + Glycerol) media. Cell mass production in different treatments were compared with the standard BL21 strain based on optical absorption at 600 nm.
Results: In the LB complex medium, Escherichia coli mutants (W3866 and W4040) grew faster than BL21 (approximately 5-fold at 8 and 10 h and 3 times at 12 h compared to samples with the dehydrogenase formate gene). However, this difference was more pronounced in the simple M9 medium, as we observed more than 6-fold growth in 24 hours of incubation in mutant samples lacking the FDH gene compared to the BL21 maternal strain.
Discussion: The experiments were completely in line with metabolic predictions and the growth of mutant bacteria was higher than that of BL21. Interestingly, most mutants grew in a simple medium containing glycerol, which showed that glycerol is a very good source for the growth of Escherichia coli bacteria. These results explain the inconsistency of predictions of previous metabolic models that declared glycerol a suitable carbon source for the growth of E. coli, but did not achieve it in practice.
Conclusion: Under normal physiological conditions, E. coli is not able to grow high in glycerol medium. Deletion of formate dehydrogenase gene caused fundamental changes in metabolic process and increased growth rate compared to BL21 strain, which indicates the inhibitory role of this enzyme in increased growth efficiency of E. coli in the presence of glycerol. In addition, the resulting strain can be used to express recombinant proteins with higher efficiency
Keywords: Escherichia coli, Metabolic pathways, Formate dehydrogenase, Increased growth rate, Iau Science.
Type of Study: Research Article |
Subject:
Microbiology
Received: 2021/12/4 | Accepted: 2021/12/31 | Published: 2022/04/11
Received: 2021/12/4 | Accepted: 2021/12/31 | Published: 2022/04/11
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