Translation Efficiency of Antiterminator Proteins Is a Determinant for the Difference in Glucose Repression of Two β-Glucoside Phosphotransferase System Gene Clusters inCorynebacterium glutamicumR

Abstract

Corynebacterium glutamicum R has two ␤-glucoside phosphoenolpyruvate, carbohydrate phosphotransferase systems (PTS) encoded by bglF and bglF2 located in the respective clusters, bglF-bglA-bglG and bglF2-bglA2-bglG2. Previously, we reported that whereas ␤-glucoside-dependent induction of bglF is strongly repressed by glucose, glucose repression of bglF2 is very weak. Here, we reveal the mechanism behind the different effects of glucose on the two bgl genes. Deletion of the ribonucleic antiterminator sequence and transcriptional terminator located upstream of the translation initiation codon of bglF markedly relieved the glucose repression of a bglF-lacZ fusion, indicating that glucose affects the antitermination mechanism that is responsible for the ␤-glucoside-dependent induction of the bglF cluster. The glucose repression of bglF mRNA was also relieved by introducing a multicopy plasmid carrying the bglG gene encoding an antiterminator of the bglF cluster. Moreover, replacement of the GUG translation initiation codon of bglG with AUG was effective in relieving the glucose repression of bglF and bglG. Inversely, expression of bglF2 and bglG2 was subject to strict glucose repression in a mutant strain in which the AUG translation initiation codon of bglG2 encoding antiterminator of the bglF2 cluster was replaced with GUG. These results suggest that the translation initiation efficiency of the antiterminator proteins, at least in part, determines whether the target genes are subject to glucose repression. We also found that bglF expression was induced by glucose in the BglG-overexpressing strains, which may be explained by the ability of BglF to transport glucose.In many bacterial cells, ␤-glucosides are taken up via the phosphoenolpyruvate:carbohydrate phosphotransferase system (PTS), which is consequently important for the acquisition of various carbohydrates (22, 34). The PTS consists of two common cytoplasmic proteins, enzyme I (EI) and HPr, and an array of carbohydrate-specific enzyme II (EII) permeases. The phosphoryl group from phosphoenolpyruvate is sequentially transferred to EI, HPr, EII, and finally to the carbohydrate as it is translocated across the membrane.An antitermination mechanism has a pivotal role in regulation of ␤-glucoside PTS genes in many bacteria. The regulation has been well studied in Escherichia coli and Bacillus subtilis. In the antitermination regulatory mechanism, transcription stops at a terminator sequence present in the 5Ј untranslated region (UTR) of the target mRNA. In the presence of substrate sugar, a transcriptional antiterminator protein of the BglG/ SacY family binds to the ribonucleic antiterminator (RAT) sequence partially overlapping the transcriptional terminator and thereby results in transcription elongation to the end of the target gene (1,5,38).An antiterminator such as E. coli BglG or B. subtilis LicT is inactivated by phosphorylation at the PTS regulation domain 1 (PRD-1) by a ␤-glucoside EII. Concomitant with PTS-dependent uptake and phosphorylation of the subs...

A unique gene cluster for the utilization of the mucosal and human milk‐associated glycans galacto‐N‐biose and lacto‐N‐biose in Lactobacillus casei

“…Although most PTS are specific for a particular carbohydrate, it has previously been described that PTS permeases transport two or more structurally related β-glucosides (Le Coq et al, 1995;Deutscher et al, 2006;Tanaka et al, 2011). While studying the metabolism of other mucosal associated glycans in L. casei we showed that fucosyl-α1,3-GlcNAc is transported by a specific PTS in a phosphorylation-independent manner before being intracellularly hydrolysed by an α-L-fucosidase (Rodriguez-Diaz et al, 2012a).…”

A third glucose uptake bypass in Corynebacterium glutamicum ATCC 31833

“…2). Genome sequencing also revealed that the bglF gene was located in the cluster bglFbglA-bglG, which has previously been identified as the β-glucoside utilization system in another wild-type C. glutamicum R (Kotrba et al 2003;Tanaka et al 2009Tanaka et al , 2011. Interestingly, a search for C. glutamicum genome databases revealed that the bglF-bglA-bglG DNA region including the upstream mutation site was missing from the corresponding position in the genome of C. glutamicum R and ATCC 13032 (Fig.…”

“…Disruption of any of the genes in strain ATCC 13032 results in a phenotype exhibiting little growth on glucose (Parche et al 2001). Exceptionally, another wildtype strain C. glutamicum R can consume glucose not only via the glucose-PTS but also via the β-glucoside-PTS because C. glutamicum R, unlike C. glutamicum ATCC 13032, has two sets of β-glucoside-PTSs that are redundantly responsible for the uptake of the β-glucosides, such as salicin and arbutin, and also glucose (Kotrba et al 2003;Tanaka et al 2009Tanaka et al , 2011.…”

Genome-Wide Analysis of the Role of Global Transcriptional Regulator GntR1 in Corynebacterium glutamicum

“…Decreased glucose utilization may relieve the catabolite repression by glucose. For example, bglF-bglA-bglG genes, coding for the ␤-glucoside catabolic genes, are under strict control of glucose repression (20). Therefore, increased expression of the bglF-bglA-bglG genes may be the result of weakened glucose repression.…”