The glgB-encoded glycogen branching enzyme is essential for glycogen accumulation in Corynebacterium glutamicum.

Corynebacterium glutamicum transiently accumulates glycogen as carbon capacitor during the early exponential growth phase in media containing carbohydrates. In some bacteria glycogen is synthesized by the consecutive action of ADP-glucose pyrophosphorylase (GlgC), glycogen synthase (GlgA) and glycogen branching enzyme (GlgB). GlgC and GlgA of C. glutamicum have been shown to be necessary for glycogen accumulation in this organism. However, although cg1381 has been annotated as the putative C. glutamicum glgB gene, cg1381 and its gene product have not been characterized and their role in transient glycogen accumulation has not yet been investigated. We show here that the cg1381 gene product of C. glutamicum catalyses the formation of α-1,6-glycosidic bonds in polysaccharides and thus represents a glycogen branching enzyme. RT-PCR experiments revealed glgB to be co-transcribed with glgE, probably encoding a maltosyltransferase. Promoter activity assays with the glgE promoter region revealed carbon-source-dependent expression of the glgEB operon. Characterization of the growth and glycogen content of glgB-deficient and glgB-overexpressing strains showed that the glycogen branching enzyme GlgB is essential for glycogen formation in C. glutamicum. Taken together these results suggest that an interplay of the enzymes GlgC, GlgA and GlgB is not essential for growth, but is required for synthesis of the transient carbon capacitor glycogen in C. glutamicum.

Glycogen formation in Corynebacterium glutamicum and role of ADP-glucose pyrophosphorylase.

Glycogen is generally assumed to serve as a major reserve polysaccharide in bacteria. In this work, glycogen accumulation in the amino acid producer Corynebacterium glutamicum was characterized, expression of the C. glutamicum glgC gene, encoding the key enzyme in glycogen synthesis, ADP-glucose (ADP-Glc) pyrophosphorylase, was analysed, and the relevance of this enzyme for growth, survival, amino acid production and osmoprotection was investigated. C. glutamicum cells grown in medium containing the glycolytic substrates glucose, sucrose or fructose showed rapid glycogen accumulation (up to 90 mg per g dry weight) in the early exponential growth phase and degradation of the polymer when the sugar became limiting. In contrast, no glycogen was detected in cells grown on the gluconeogenic substrates acetate or lactate. In accordance with these results, the specific activity of ADP-Glc pyrophosphorylase was 20-fold higher in glucose-grown than in acetate- or lactate-grown cells. Expression analysis suggested that this carbon-source-dependent regulation might be only partly due to transcriptional control of the glgC gene. Inactivation of the chromosomal glgC gene led to the absence of ADP-Glc pyrophosphorylase activity, to a complete loss of intracellular glycogen in all media tested and to a distinct lag phase when the cells were inoculated in minimal medium containing 750 mM sodium chloride. However, the growth of C. glutamicum, its survival in the stationary phase and its glutamate and lysine production were not affected by glgC inactivation under either condition tested. These results indicate that intracellular glycogen formation is not essential for growth and survival of and amino acid production by C. glutamicum and that ADP-Glc pyrophosphorylase activity might be advantageous for fast adaptation of C. glutamicum to hyperosmotic stress.