Functional characterization of sucrose phosphorylase and scrR, a regulator of sucrose metabolism in Lactobacillus reuteri

Abstract

Lactobacillus reuteri harbours alternative enzymes for sucrose metabolism, sucrose phosphorylase, fructansucrases, and glucansucrases. Sucrose phosphorylase and fructansucrases additionally contribute to raffinose metabolism. Glucansucrases and fructansucrases produce exopolysaccharides as alternative to sucrose hydrolysis. L. reuteri LTH5448 expresses a levansucrase (ftfA) and sucrose phosphorylase (scrP), both are inducible by sucrose. This study determined the contribution of scrP to sucrose and raffinose metabolism in L. reuteri LTH5448, and elucidated the role of scrR in regulation sucrose metabolism. Disruption of scrP and scrR was achieved by double crossover mutagenesis. L. reuteri LTH5448, LTH5448ΔscrP and LTH5448ΔscrR were characterized with respect to growth and metabolite formation with glucose, sucrose, or raffinose as sole carbon source. Inactivation of scrR led to constitutive transcription of scrP and ftfA, demonstrating that scrR is negative regulator. L. reuteri LTH5448 and the LTH5448ΔscrP or LTH5448ΔscrR mutant strains did not differ with respect to glucose, sucrose or raffinose utilization. However, L. reuteri LTH5448ΔscrP produced more levan, indicating that the lack of sucrose phosphorylase is compensated by an increased metabolic flux through levansucrase. In conclusion, the presence of alternate pathways for sucrose and raffinose metabolism and their regulation indicate that these substrates, which are abundant in plants, are preferred carbohydrate sources for L. reuteri.

Keywords: FOS; FtfA; GI; HPAEC–PAD; HPLC; LAB; LB; Lactobacillus; Lactobacillus reuteri; LevS; Luria–Bertani; MRS; PTS; ScrP; ScrR; Sourdough; Sucrose; Sucrose phosphorylase; deMan–Rogosa–Sharpe medium; fructo-oligosaccharides; fructosyltransferase A; gastrointestinal; high performance anion exchange chromatography with integrated pulsed amperometric detection; high performance liquid chromatography; lactic acid bacteria; levansucrase; phosphotransferase system; qPCR; quantitative real time polymerase chain reaction; sucrose phosphorylase; sucrose regulator.