Meekrathok Piyanat, Bürger Marco, Porfetye Arthur T, Vetter Ingrid R, Suginta Wipa
Biochemistry-Electrochemistry Research Unit, School of Biochemistry, Institute of Science, Suranaree University of Technology, Nakhon Ratchasima 30000, Thailand.
Max Planck Institute of Molecular Physiology, 44227 Dortmund, Germany.
Acta Crystallogr F Struct Biol Commun. 2015 Apr;71(Pt 4):427-33. doi: 10.1107/S2053230X1500415X. Epub 2015 Mar 20.
Vibrio harveyi β-N-acetylglucosaminidase (VhGlcNAcase) is a new member of the GH20 glycoside hydrolase family responsible for the complete degradation of chitin fragments, with N-acetylglucosamine (GlcNAc) monomers as the final products. In this study, the crystallization and preliminary crystallographic data of wild-type VhGlcNAcase and its catalytically inactive mutant D437A in the absence and the presence of substrate are reported. Crystals of wild-type VhGlcNAcase were grown in 0.1 M sodium acetate pH 4.6, 1.4 M sodium malonate, while crystals of the D437A mutant were obtained in 0.1 M bis-tris pH 7.5, 0.1 M sodium acetate, 20% PEG 3350. X-ray data from the wild-type and the mutant crystals were collected at a synchrotron-radiation light source and were complete to a resolution of 2.5 Å. All crystals were composed of the same type of dimer, with the substrate N,N'-diacetylglucosamine (GlcNAc₂ or diNAG) used for soaking was cleaved by the active enzyme, leaving only a single GlcNAc molecule bound to the protein.
哈维氏弧菌β-N-乙酰氨基葡萄糖苷酶（VhGlcNAcase）是GH20糖苷水解酶家族的新成员，负责将几丁质片段完全降解，最终产物为N-乙酰氨基葡萄糖（GlcNAc）单体。在本研究中，报道了野生型VhGlcNAcase及其催化无活性突变体D437A在有无底物情况下的结晶和初步晶体学数据。野生型VhGlcNAcase的晶体在0.1 M醋酸钠pH 4.6、1.4 M丙二酸钠中生长，而D437A突变体的晶体在0.1 M双三羟甲基氨基甲烷pH 7.5、0.1 M醋酸钠、20%聚乙二醇3350中获得。野生型和突变体晶体的X射线数据在同步辐射光源处收集，完整分辨率达到2.5 Å。所有晶体均由同一种二聚体组成，用于浸泡的底物N,N'-二乙酰氨基葡萄糖（GlcNAc₂或二NAG）被活性酶切割，仅留下一个与蛋白质结合的单个GlcNAc分子。
