Investigation of storage polysaccharide metabolism in lactic acid bacteria

Abstract

Glycogen biosynthesis and its regulation have been investigated extensively in bacteria at the level of ADP-Glc synthesis. The enzyme ADP-glucose pyrophosphorylase (ATP: α-dglucose-1-phosphate adenylyltransferase; EC 2.7.7.27; ADP-Glc PPase) catalyzes ADP-Glc formation in a reversible reaction which is the rate determining step of glycogen biosynthesis. This enzyme is encoded by glgC in all bacteria. However, the role of an additional, similar gene glgD in Gram-positive bacteria including lactic acid bacteria (LAB) is poorly understood. Only a regulatory role of glgD has been reported in Bacillus stearothermophilus without any activity being shown for the protein product GlgD. Therefore, it was decided to investigate the function of glgC-homologous to glgD gene in LAB. The focused new role of glgD might reveal the mechanism of regulation of glycogen in this group of bacteria. And possibility link intracellular polysaccharide (IPS) formation popular probiotic characterisation of certain species of LAB. Our studies encompassed the LAB which are a group of facultative anaerobic, non-pathogenic, non-colonizing, non sporulating Gram-positive bacteria. The genes for glycogen synthesis have been found in the operon glgBCDAP or glgCDAP-B as in Lactobacillus plantarum WCFS1 and Lactococcus lactis subsp. cremoris MG1363, respectively. We have focused on the detailed functional analysis of both the genes glgC and glgD in L. lactis subsp cremoris MG1363 and SK11 and Lb. plantarum WCFS1. In a fusion expression study of both the genes glgC and glgD from Lb. plantarum WCFS1, we have shown experimentally that the proteins GlgC and GlgD have interact with each other. Both the proteins appear to be subunits forming the fully active enzyme with an α/β heteroligomer type structure. To our knowledge, this is the first study to prove that the GlgC & GlgD proteins have an interaction in lactic acid bacteria. Moreover, we have also observed a low enzymatic activity of the the GlgD protein in presence of ATP and a high affinity for UTP. The first observation suggests that GlgD has ADP-Glc-PPase activity in addition to GlgC and the second is indicative of an alternative reaction to produce UDP-Glucose possibly providing to an alternate pathway for glycogen biosynthesis.