N-酰基葡糖胺衍生物对GABA转运体功能的影响

为筛选GAT蛋白活性抑制剂，以稳定表达GAT1蛋白的人胚胎肾细胞(HEK-293)为模型，利用GABA摄取定量测试测定N-酰基葡糖胺衍生物对该蛋白功能的影响。同时，由于GAT1蛋白的功能与其N-糖链结构尤其是末端唾液酸密切相关，再通过定量该蛋白糖链末端唾液酸的表达分析衍生物对蛋白糖链修饰的作用。结果显示，10 mmol/L 3-O-甲基-N-乙酰基葡糖胺(3-O-Met-GlcNAc)通过抑制GAT1蛋白的唾液酸化(降至66.8%)将GABA摄取活性减弱到到53%(P<；0.01)；而10 mmol/L N-丙酰基葡糖胺(GlcNProp)，N-环丙甲酰基葡糖胺(GlcNCyclo)，N-己酰基葡糖胺(GlcNHex)，N-乙酰氨基乙酰基葡糖(GlcNAc-acetamido)则通过抑制蛋白糖链末端修饰分别将GABA摄取活性减弱到54%、63%、63%和67%(P<；0.01)。因此，N-酰基葡糖胺衍生物可用于进一步筛选研究GAT蛋白活性抑制剂或唾液酸生物合成抑制剂。

Effects of N-acyl glucosamines on GABA uptake activity of GABA transporter 1

In order to screen the inhibitors of GAT1 protein, the effects of synthetic N-acylglucosamines on the GABA uptake activity of GAT1 were examined in the HEK-293 cell model stably expressing GAT1 by performing GABA uptake assay. And since the N-linked oligosaccharides, especially their terminal sialic acid of GAT1 are necessary for the function of GAT1, their effects on the modification of N-glycans of GAT1 were also determined by the quantitative analysis of the sialic acid of GAT1. The results showed that 10 mmol /L 3-O-methyl-N-acetylglucosamine(3-O-Met-GlcNAc)reduced the GABA uptake activity of GAT1 to 53%(P< 0. 01)through inhibiting the sialylation of GAT1(66. 8%). And the GABA uptake activities were decreased to 54%, 63%, 63% and 67%(P< 0. 01), respectively, by the treatments of 10 mmol /L N-propionylglucosamine(GlcNProp), N-hexanoylglucosamine(GlcNHex), N-cycloproylgormylglucosamine(GlcNCyclo)and N-acetamidoacetylglucosamine(GlcNAc-acetamido)through inhibiting N-glycan trimming. These results indicate that the analogues of glucosamines have great potential in the development of inhibitors of GAT1 activity or sialic acid biosynthesis.