Corynebacterium glutamicum is an excellent protein expression host. Although the entire genome of C. glutamicum has been sequenced, some key genes that affect protein expression have not yet been researched. The gene NCgl2632 is predicted to be one of the key genes affecting the energy metabolism of C. glutamicum. Therefore, the NCgl2632 knockout strain and NCgl2632 overexpression strain were constructed, and the growth curve and changes of intracellular ATP levels were measured. Further, enhanced green fluorescent protein (eGFP) and variable domain of heavy chain antibody (VHH) were used to verify the effect of changes in the gene expression level on the expression of exogenous proteins, which indicated that knocking out the gene improved the expression of exogenous proteins. At the same time, since the knockout phenotype of NCgl0909 was proved to increase the expression of exogenous proteins in previous work, the C.g 15647ΔNCgl2632ΔNCgl0909 co-knockout strain was constructed on this basis. The model protein VHH was used to validate the ability of expressing exogenous proteins in the co-knockout strain. It was proved that the co-knockout strain was beneficial to the expression of VHH. In addition, a potential protein human MutT homolog 1(hMTH1) was selected for expression verification and activity detection, in which case the co-knockout strain also increased the expression of hMTH1. Owing to its small molecular weight and stability, hMTH1 can be used as a new model protein to verify the ability of strains to produce exogenous proteins in the future. Finally, to obtain a better medium ratio for protein expression, the fermentation medium of the co-knockout strain was optimized.
ZHANG Qi
,
LIU Xiuxia
,
YANG Yankun
,
BAI Zhonghu
. Effect of knockout and overexpression of gene NCgl2632 on expression of three exogenous proteins in Corynebacterium glutamicum[J]. Food and Fermentation Industries, 2022
, 48(14)
: 1
-8
.
DOI: 10.13995/j.cnki.11-1802/ts.029769
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