Medical Proteomics Research Center, Korea Institute of Bioscience and Biotechnology, Daejeon, Korea.
Proteins. 2010 Jun;78(8):1847-55. doi: 10.1002/prot.22697.
Glycogen serves as major energy storage in most living organisms. GlgX, with its gene in the glycogen degradation operon, functions in glycogen catabolism by selectively catalyzing the debranching of polysaccharide outer chains in bacterial glycosynthesis. GlgX hydrolyzes alpha-1,6-glycosidic linkages of phosphorylase-limit dextrin containing only three or four glucose subunits produced by glycogen phosphorylase. To understand its mechanism and unique substrate specificity toward short branched alpha-polyglucans, we determined the structure of GlgX from Escherichia Coli K12 at 2.25 A resolution. The structure reveals a monomer consisting of three major domains with high structural similarity to the subunit of TreX, the oligomeric bifunctional glycogen debranching enzyme (GDE) from Sulfolobus. In the overlapping substrate binding groove, conserved residues Leu270, Asp271, and Pro208 block the cleft, yielding a shorter narrow GlgX cleft compared to that of TreX. Residues 207-213 form a unique helical conformation that is observed in both GlgX and TreX, possibly distinguishing GDEs from isoamylases and pullulanases. The structural feature observed at the substrate binding groove provides a molecular explanation for the unique substrate specificity of GlgX for G4 phosphorylase-limit dextrin and the discriminative activity of TreX and GlgX toward substrates of varying lengths.
糖原是大多数生物体内主要的储能物质。GlgX 及其基因位于糖原降解操纵子中，通过选择性催化多糖外链的分支酶解来参与糖原分解代谢。GlgX 水解糖原磷酸化酶产生的仅含有三到四个葡萄糖单元的磷酸化酶限制糊精中的α-1,6-糖苷键，这些糊精是由糖原磷酸化酶产生的。为了了解其机制和对短分支α-多聚糖的独特底物特异性，我们解析了大肠杆菌 K12 的 GlgX 结构，分辨率为 2.25Å。结构显示单体由三个主要结构域组成，与 TreX 的亚基具有高度结构相似性，TreX 是来自 Sulfolobus 的寡聚多功能糖原分支酶（GDE）。在重叠的底物结合槽中，保守残基 Leu270、Asp271 和 Pro208 阻塞了裂缝，导致 GlgX 的裂缝比 TreX 更短更窄。残基 207-213 形成独特的螺旋构象，在 GlgX 和 TreX 中都观察到，可能将 GDE 与异淀粉酶和普鲁兰酶区分开来。在底物结合槽观察到的结构特征为 GlgX 对 G4 磷酸化酶限制糊精的独特底物特异性以及 TreX 和 GlgX 对不同长度底物的区分活性提供了分子解释。
