Maltose Uptake by the Novel ABC Transport System MusEFGK ₂ I Causes Increased Expression of ptsG in Corynebacterium glutamicum

Abstract

The Gram-positive Corynebacterium glutamicum efficiently metabolizes maltose by a pathway involving maltodextrin and glucose formation by 4-␣-glucanotransferase, glucose phosphorylation by glucose kinases, and maltodextrin degradation via maltodextrin phosphorylase and ␣-phosphoglucomutase. However, maltose uptake in C. glutamicum has not been investigated. Interestingly, the presence of maltose in the medium causes increased expression of ptsG in C. glutamicum by an unknown mechanism, although the ptsG-encoded glucose-specific EII permease of the phosphotransferase system itself is not required for maltose utilization. We identified the maltose uptake system as an ABC transporter encoded by musK (cg2708; ATPase subunit), musE (cg2705; substrate binding protein), musF (cg2704; permease), and musG (cg2703; permease) by combination of data obtained from characterization of maltose uptake and reanalyses of transcriptome data. Deletion of the mus gene cluster in C. glutamicum ⌬mus abolished maltose uptake and utilization. Northern blotting and reverse transcription-PCR experiments revealed that musK and musE are transcribed monocistronically, whereas musF and musG are part of an operon together with cg2701 (musI), which encodes a membrane protein of unknown function with no homologies to characterized proteins. Characterization of growth and [14 C]maltose uptake in the musI insertion strain C. glutamicum IMcg2701 showed that musI encodes a novel essential component of the maltose ABC transporter of C. glutamicum. Finally, ptsG expression during cultivation on different carbon sources was analyzed in the maltose uptake-deficient strain C. glutamicum ⌬mus. Indeed, maltose uptake by the novel ABC transport system MusEFGK 2 I is required for the positive effect of maltose on ptsG expression in C. glutamicum.

Transcription of Sialic Acid Catabolism Genes in Corynebacterium glutamicum Is Subject to Catabolite Repression and Control by the Transcriptional Repressor NanR

“…Slot blot experiments were performed as described previously (60). For hybridization, digoxigenin (DIG)-11-dUTP-labeled gene-specific antisense RNA probes against ptsG mRNA and the 16S rRNA were prepared from PCR products (generated with oligonucleotides listed in Table S2) carrying the T7 promoter by in vitro transcription (1 h at 37°C) using T7 RNA polymerase (MBI Fermentas) as described previously (61).…”

The α-Glucan Phosphorylase MalP of Corynebacterium glutamicum Is Subject to Transcriptional Regulation and Competitive Inhibition by ADP-Glucose

“…However, malP transcript levels, as well as MalP activities, increased in the bioreactor experiments already at a residual maltose concentration of ϳ32 mM ( Fig. 2A), which is 1,550-fold higher than the maltose concentration of ϳ20 M required for the maltose uptake system MusEFGK 2 I to work at its maximal velocity (38). Thus, the increase in malP transcription sets actually in before the substrate becomes limiting and it seems that C. glutamicum anticipates the start of substrate limitation and starts to change its metabolism in preparation for the stationary growth phase.…”

“…Indeed, two genes for ␣-glucan phosphorylases reside in the C. glutamicum genome, namely, glgP1 (cg1479) and glgP2 (cg2289) (37). The metabolic pathway for maltose metabolism in C. glutamicum is similar to the pathway known from E. coli and involves maltose uptake by the ABC transporter MusEFGK 2 I, maltodextrin formation by MalQ, degradation of maltodextrins to glucose-1-phosphate (glc-1-P) by an ␣-glucan phosphorylase, and glucose-6-phosphate (glc-6-P) formation by glucokinases and phosphoglucomutases (38)(39)(40)(41). Based on the absence of maltodextin phosphorylase activity in the glgP1 deletion mutant strain and the severe growth phenotype of this strain observed exclusively in cultivations on maltose, the glgP1 gene product was assigned the function of a maltodextrin phosphorylase and the gene therefore was renamed malP (41).…”

“…The presence of mechanisms for the control of maltose metabolism, as well as for the coordination of glycogen synthesis and degradation, is expected for C. glutamicum; however, only the transcriptional control of genes for the maltose uptake system and of the genes glgC and glgA required for the glycogen synthesis have been investigated thus far (38,44). Mechanisms for the control of enzyme activities involved in either glycogen degradation or maltose metabolism have not yet been analyzed in C. glutamicum.…”

Transcription of malP is subject to phosphotransferase system-dependent regulation in Corynebacterium glutamicum

“…Maltose uptake studies were performed essentially as described by Henrich et al (2013). In detail, C. glutamicum cells were grown in the media indicated in the text to mid-exponential growth phase, harvested by centrifugation, washed twice with ice-cold CGC medium, suspended to an OD 600 of 2 with CGC medium and stored on ice until the measurement was carried out.…”