Identification and application of a novel strong constitutive promoter in Corynebacterium glutamicum

The replacement of promoters with various strengths is an effective strategy to fine-tune gene expression. More available promoters with a broad range of transcription efficiency are needed in metabolic engineering. In this study, a putative protein coding gene CP_2454 was identified with a stable and high transcriptional level from an l-leucine-producing strain Corynebacterium glutamicum CP, which was absent in wild-type C. glutamicum ATCC 13032. The transcriptional level of CP_2454 was about 80.0% those of tuf and sod, and was 1.6 and 3.2 times those of ilvB and gapA, respectively. These promoters were cloned into C. glutamicum ATCC 13032 to control the expression of green fluorescent protein (GFP). While, the expressional level of GFP under the control of PCP_2454 was close to that of Ptuf and Psod, which was significantly higher than that of other tested promoters. The native promoter of ilvB controlling the expression of a feedback-resistant acetolactate synthase was replaced by PCP_2454, resulting in an increase of l-valine titer by 58.5%. A further 24.9% increase of l-valine titer was achieved after replacement of the promoter of ilvD encoding dihydroxyacid dehydratase with PCP_2454. Identification of the constitutive promoter PCP_2454 expands the promoter library of C. glutamicum and provides essential reference for tunable expression of target genes in C. glutamicum.