Xiuling Shang1, Xin Chai2,3, Xuemei Lu2,3, Yuan Li2,3, Yun Zhang1, Guoqiang Wang2,3, Chen Zhang2,3, Shuwen Liu1, Yu Zhang1, Jiyin Ma2,3, Tingyi Wen4,5,6. 1. CAS Key Laboratory of Pathogenic Microbiology and Immunology, Institute of Microbiology, Chinese Academy of Sciences, 1 Beichen West Road, Chaoyang District, Beijing, 100101, China. 2. Beijing Zhongke EPPEN Biotechnology Co., Ltd, Beijing, 100085, China. 3. Ningxia EPPEN Biotechnology Co., Ltd, Yongning, 750100, Ningxia, China. 4. CAS Key Laboratory of Pathogenic Microbiology and Immunology, Institute of Microbiology, Chinese Academy of Sciences, 1 Beichen West Road, Chaoyang District, Beijing, 100101, China. wenty@im.ac.cn. 5. Beijing Zhongke EPPEN Biotechnology Co., Ltd, Beijing, 100085, China. wenty@im.ac.cn. 6. Savaid Medical School, University of Chinese Academy of Sciences, Beijing, 100049, China. wenty@im.ac.cn.
Abstract
OBJECTIVE: To identify useful native promoters of Corynebacterium glutamicum for fine-tuning of gene expression in metabolic engineering. RESULTS: Sixteen native promoters of C. glutamicum were characterized. These promoters covered a strength range of 31-fold with small increments and exhibited relatively stable activity during the whole growth phase using β-galactosidase as the reporter. The mRNA level and enzymatic activity of the lacZ reporter gene exhibited high correlation (R 2 = 0.96) under the control of these promoters. Sequence analysis found that strong promoters had high similarity of the -10 hexamer to the consensus sequence and preference of the AT-rich UP element upstream the -35 region. To test the utility of the promoter library, the characterized native promoters were applied to modulate the sucCD-encoded succinyl-CoA synthetase expression for L-lysine overproduction. CONCLUSIONS: The native promoters with various strengths realize the efficient and precise regulation of gene expression in metabolic engineering of C. glutamicum.
OBJECTIVE: To identify useful native promoters of Corynebacterium glutamicum for fine-tuning of gene expression in metabolic engineering. RESULTS: Sixteen native promoters of C. glutamicum were characterized. These promoters covered a strength range of 31-fold with small increments and exhibited relatively stable activity during the whole growth phase using β-galactosidase as the reporter. The mRNA level and enzymatic activity of the lacZ reporter gene exhibited high correlation (R 2 = 0.96) under the control of these promoters. Sequence analysis found that strong promoters had high similarity of the -10 hexamer to the consensus sequence and preference of the AT-rich UP element upstream the -35 region. To test the utility of the promoter library, the characterized native promoters were applied to modulate the sucCD-encoded succinyl-CoA synthetase expression for L-lysine overproduction. CONCLUSIONS: The native promoters with various strengths realize the efficient and precise regulation of gene expression in metabolic engineering of C. glutamicum.