Construction of a Vitreoscilla Hemoglobin Promoter-Based Tunable Expression System for Corynebacterium glutamicum

Abstract

Corynebacterium glutamicum is an industrial strain used for the production of valuable chemicals such as L-lysine and L-glutamate. Although C. glutamicum has various industrial applications, a limited number of tunable systems are available to engineer it for efficient production of platform chemicals. Therefore, in this study, we developed a novel tunable promoter system based on repeats of the Vitreoscilla hemoglobin promoter (P-vgb). Tunable expression of green fluorescent protein (GFP) was investigated under one, four, and eight repeats of P-vgb (P-vgb, P-vgb4, and P-vgb8). The intensity of fluorescence in recombinant C. glutamicum strains increased as the number of P-vgb increased from single to eight (P-vgb8) repeats. Furthermore, we demonstrated the application of the new P-vgb promoter-based vector system as a platform for metabolic engineering of C. glutamicum by investigating 5-aminovaleric acid (5-AVA) and gamma-aminobutyric acid (GABA) production in several C. glutamicum strains. The profile of 5-AVA and GABA production by the recombinant strains were evaluated to investigate the tunable expression of key enzymes such as DavBA and GadB(mut). We observed that 5-AVA and GABA production by the recombinant strains increased as the number of P-vgb used for the expression of key proteins increased. The recombinant C. glutamicum strain expressing DavBA could produce higher amounts of 5-AVA under the control of P-vgb8 (3.69 +/- 0.07 g/L) than the one under the control of P-vgb (3.43 +/- 0.10 g/L). The average gamma-aminobutyric acid production also increased in all the tested strains as the number of P-vgb used for GadB(mut) expression increased from single (4.81-5.31 g/L) to eight repeats (4.94-5.58 g/L).