LactoSpanks: a collection of IPTG inducible promoters for the commensal lactic acid bacteriaLactobacillus gasseri

Abstract

ABSTRACTLactic acid bacteria (LAB) are important for many biotechnological applications, such as bioproduction and engineered probiotics for therapy. Inducible promoters are key gene expression control elements, yet those available in LAB are mainly based on bacteriocin systems and have many drawbacks, including large gene clusters, costly inducer peptides and little portability toin vivosettings. UsingLactobacillus gasseri, a model commensal bacteria from the human gut, we report the engineering of Lactospanks promoters (Pls), a collection of variable strength inducible promoters controlled by theLacIrepressor fromB. subtilisand induced by isopropyl β-D-1-thiogalactopyranoside (IPTG). We first show that the Phyper-spank promoter fromBacillus subtilisis functional inL. gasseri, albeit with substantial leakage. We then construct and screen a semi-rational library of Phyper-spank variants to select a set of four IPTG-inducible promoters that span a range of expression levels and exhibit reduced leakages and operational dynamic ranges (from ca. 9 to 28 fold-change). With their low genetic footprint and simplicity of use, Lactospanks will support many applications inL. gasseri, and potentially other lactic acid and gram-positive bacteria.