Rapid Generation of Universal Synthetic Promoters for Controlled Gene Expression in Both Gas-Fermenting and Saccharolytic <i>Clostridium</i> Species

Abstract

Engineering solventogenic clostridia, a group of important industrial microorganisms, to realize their full potential in biorefinery application is still hindered by the absence of plentiful biological parts. Here, we developed an effective approach for rapid generation of a synthetic promoter library in solventogenic clostridia based on a dual-reporter system (catP-lacZ) and a widely used strong thl promoter. The yielded artificial promoters, spanning 2 orders of magnitude, comprised two modular components (the core promoter region and the spacer between RBS and the translation-initiating code), and the strongest promoter had an over 10-fold-higher activity than the original expression part P_thl. The test of these synthetic promoters in controlled expression of sadh and danK in saccharolytic C. acetobutylicum and gas-fermenting C. ljungdahlii, respectively, gave the expected phenotypes, and moreover, showed good correlation between promoter activities and phenotypic changes. The presented wide-strength-range promoters here will be useful for synthetic biology application in solventogenic clostridia.