Biological significance of N-acetylglucosamine 6-sulfation

表达3种N-乙酰氨基葡萄糖-6-磺基转移酶（GlcNAc 6ST），即GlcNAc 6ST-1、GlcNAc 6ST-2、GlcNAc 6ST-3，并比较其特异性。GlcNAc 6ST-1和GlcNAc 6ST-2作用于粘蛋白型核心2糖（GlcNAcβ1-6[Galβ1-3]GalNAc），并且两种酶之间没有显著差异。虽然已知GlcNAc 6ST-2参与淋巴细胞归巢，但尚不清楚GlcNAc 6ST-1是否发挥这种作用。用GlcNAc 6ST-1和岩藻糖基转移酶VII双重转染的细胞诱导表达L-选择素的细胞滚动。GlcNAc 6ST-3仅作用于核心2糖。另一方面，只有GlcNAc 6ST-2作用于核心3糖（GlcNAcβ1-3 GalNAc）。根据该特异性，认为GlcNAc 6ST-2是在粘液腺癌中新表达的酶，通过RT-PCR分析确定了这一点。通过Southern和北方印迹分析证实该小鼠完全缺乏GlcNAc 6ST-1。小鼠正常出生，没有表现出明显的异常。目前正在进行回交，因此我们能够分析淋巴细胞归巢中可能的异常。使用杆状病毒系统在Tn 5细胞中产生GilcNAc 6ST-1作为融合蛋白。纯化的重组酶可用于合成硫酸化寡糖和糖蛋白结合糖的硫酸化。

Three species of N-acetylglucosamine-6-sulfotransferases (GlcNAc6ST), namely GlcNAc6ST-1, GlcNAc6ST-2, GlcNAc6ST-3 were expressed and the specificities were compared. GlcNAc6ST-l as well as GlcNAc6ST-2 acted on mucin-type core 2 sugar (GlcNAcβ1-6[Galβ1-3]GalNAc) and there were not significant differences between the two enzymes. Although GlcNAc6ST-2 is known to be involved in lymphocyte homing, it is not clarified whether GlcNAc6ST-1 plays such a role. Cells doubly transfected with GlcNAc6ST-1 and fucosyltransferase VII induced roling of L-selectin-expressing cells. GlcNAc6ST-3 acted only on core 2 sugars. On the other hand, only GlcNAc6ST-2 acted on core 3 sugars (GlcNAcβ1-3 GalNAc). From this specificity, GlcNAc6ST-2 was considered to be the enzyme, which became newly expressed in mucinous adenocarcinoma, and this point was confined by RT-PCR analysis.We produced knockout mice deficient in the GlcNAc6ST-1 gene. That this mice completely lacked GlcNAc6ST-1 was confirmed by Southern and Northern blot analyses. The mice was born normally, and showed no apparent abnormality. Currently backcross is on the way, so that we are able to analyze possible abnormality in lymphocyte homing.GilcNAc6ST-1 was produced as fasion protein in Tn5 cells using the Baculovirus system. The purified recombinant enzyme was useful for synthesis of sulfated oligosaccharides and sulfation of glycoprotein-bound carbohydrates.