Instituto de Agrobiotecnología del Litoral (UNL-CONICET), Facultad de Bioquímica y Ciencias Biológicas, Paraje 'El Pozo' CC 242, S3000ZAA, Santa Fe, Argentina; Department of Chemistry and Biochemistry, Loyola University Chicago, 1068, W Sheridan Rd., Chicago, IL, 60660, USA.
Mol Microbiol. 2013 Dec;90(5):1011-27. doi: 10.1111/mmi.12413. Epub 2013 Oct 29.
Streptococcus mutans is the leading cause of dental caries worldwide. The bacterium accumulates a glycogen-like internal polysaccharide, which mainly contributes to its carionegic capacity. S.mutans has two genes (glgC and glgD) respectively encoding putative ADP-glucose pyrophosphorylases (ADP-Glc PPase), a key enzyme for glycogen synthesis in most bacteria. Herein, we report the molecular cloning and recombinant expression of both genes (separately or together) followed by the characterization of the respective enzymes. When expressed individually GlgC had ADP-Glc PPase activity, whereas GlgD was inactive. Interestingly, the coexpressed GlgC/GlgD protein was one order of magnitude more active than GlgC alone. Kinetic characterization of GlgC and GlgC/GlgD pointed out remarkable differences between them. Fructose-1,6-bis-phosphate activated GlgC by twofold, but had no effect on GlgC/GlgD. Conversely, phospho-enol-pyruvate and inorganic salts inhibited GlgC/GlgD without affecting GlgC. However, in the presence of fructose-1,6-bis-phosphate GlgC acquired a GlgC/GlgD-like behaviour, becoming sensitive to the stated inhibitors. Results indicate that S. mutans ADP-Glc PPase is an allosteric regulatory enzyme exhibiting sensitivity to modulation by key intermediates of carbohydrates metabolism in the cell. The particular regulatory properties of the S.mutans enzyme agree with phylogenetic analysis, where GlgC and GlgD proteins found in other Firmicutes arrange in distinctive clusters.
变形链球菌是全球范围内导致龋齿的主要原因。该细菌积累了一种类似糖原的内部多糖，主要有助于其致龋能力。变形链球菌有两个基因（glgC 和 glgD），分别编码假定的 ADP-葡萄糖焦磷酸化酶（ADP-Glc PPase），这是大多数细菌中糖原合成的关键酶。本文报道了这两个基因（分别或一起）的分子克隆和重组表达，以及各自酶的特性。当单独表达时，GlgC 具有 ADP-Glc PPase 活性，而 GlgD 则没有活性。有趣的是，共表达的 GlgC/GlgD 蛋白的活性比单独的 GlgC 高一个数量级。GlgC 和 GlgC/GlgD 的动力学特征表明它们之间存在显著差异。果糖-1,6-二磷酸使 GlgC 的活性增加了两倍，但对 GlgC/GlgD 没有影响。相反，磷酸烯醇丙酮酸和无机盐抑制 GlgC/GlgD，而不影响 GlgC。然而，在果糖-1,6-二磷酸存在下，GlgC 获得了类似于 GlgC/GlgD 的行为，对所述抑制剂变得敏感。结果表明，变形链球菌 ADP-Glc PPase 是一种变构调节酶，对细胞内碳水化合物代谢的关键中间产物的调节具有敏感性。该酶的特殊调节特性与系统发育分析一致，在其他Firmicutes 中发现的 GlgC 和 GlgD 蛋白排列在独特的簇中。
