Killivalavan Asaithambi, Zhuang Ningning, Park Young Shik, Lee Kon Ho
Department of Microbiology, School of Medicine, Gyeongsang National University, Jinju 660-751, Republic of Korea.
Plant Molecular Biology and Biotechnology Research Center (PMBBRC), Gyeongsang National University, Jinju 660-701, Republic of Korea.
Acta Crystallogr F Struct Biol Commun. 2014 Feb;70(Pt 2):203-5. doi: 10.1107/S2053230X13034298. Epub 2014 Jan 21.
A UDP-glucose:tetrahydrobiopterin α-glucosyltransferase (BGluT) enzyme was discovered in the cyanobacterium Synechococcus sp. PCC 7942 which transfers a glucose moiety from UDP-glucose to tetrahydrobiopterin (BH4). BGluT protein was overexpressed with selenomethionine labelling for structure determination by the multi-wavelength anomalous dispersion method. The BGluT protein was purified by nickel-affinity and size-exclusion chromatography. It was then crystallized by the hanging-drop vapour-diffusion method using a well solution consisting of 0.1 M bis-tris pH 5.5, 19%(w/v) polyethylene glycol 3350 with 4%(w/v) D(+)-galactose as an additive. X-ray diffraction data were collected to 1.99 Å resolution using a synchrotron-radiation source. The crystals belonged to the monoclinic space group C2, with unit-cell parameters a = 171.35, b = 77.99, c = 53.77 Å, β = 90.27°.
在蓝藻聚球藻属嗜热栖热菌（Synechococcus sp. PCC 7942）中发现了一种UDP-葡萄糖：四氢生物蝶呤α-葡萄糖基转移酶（BGluT），该酶可将UDP-葡萄糖中的葡萄糖部分转移至四氢生物蝶呤（BH4）。通过多波长反常散射法对用硒代甲硫氨酸标记的BGluT蛋白进行结构测定，该蛋白过表达。BGluT蛋白通过镍亲和色谱法和尺寸排阻色谱法进行纯化。然后，采用悬滴气相扩散法，以含有0.1 M双三羟甲基氨基甲烷（bis-tris）pH 5.5、19%（w/v）聚乙二醇3350以及4%（w/v）D（+）-半乳糖作为添加剂的储液进行结晶。使用同步辐射光源收集X射线衍射数据，分辨率达到1.99 Å。晶体属于单斜空间群C2，晶胞参数为a = 171.35、b = 77.99、c = 53.77 Å，β = 90.27°。
