Glutamine-fructose-6-phosphate aminotransferase的表达纯化以及结晶

Crystallization and Preliminary X-ray Data Collection of Glutamine-fructose-6-amidotrasperase

Glutamine-fructose-6-phosphate amidotransferase(Glm S)在己糖胺的生物合成途径中是一个发挥关键作用的限速酶.Glm S能够将fructose 6-phosphate(Fru-6P)转移到glucosamine-6P或者glucose-6P上,这个过程依赖于谷氨酰胺的呈现或者是缺失.在研究过程中,利用Ni亲和层析柱对目的蛋白进行初步纯化,再用离子柱和分子筛层析进行进一步的纯化去除标签以及杂蛋白.纯化后的目的蛋白浓缩至适宜浓度,进行晶体筛选以及优化.最后将适宜的晶体进行初步的X射线衍射分析.

Glutamine-fructose-6-amidotransferase（Glm S） is a critical rate-limiting enzyme in the biosynthetic pathway of hexosamine. Glm S transfers fructose- 6-phosphate（Fru-6P） into glucosamine-6P or glucose-6P which depends on the presence or absence of Glutamine. Here, we purified the fusion protein with Ni-NTA column firstly, and then Hitrap Q and Hi Load 26/60 Superdex 200 size-exclusion column were used to remove the his tag and other proteins. The purified protein was subjected into crystal screening and optimization. And finally qualified crystals were used for x-ray diffractions.