Department of Integrated Biosciences, Graduate School of Frontier Sciences, University of Tokyo, Chiba 277-8562, Japan.
J Biol Chem. 2010 Feb 19;285(8):5178-87. doi: 10.1074/jbc.M109.018663. Epub 2009 Dec 3.
We previously found that pigeon IgG possesses unique N-glycan structures that contain the Gal alpha1-4Gal beta1-4Gal beta1-4GlcNAc sequence at their nonreducing termini. This sequence is most likely produced by putative alpha1,4- and beta1,4-galactosyltransferases (GalTs), which are responsible for the biosynthesis of the Gal alpha1-4Gal and Gal beta1-4Gal sequences on the N-glycans, respectively. Because no such glycan structures have been found in mammalian glycoproteins, the biosynthetic enzymes that produce these glycans are likely to have distinct substrate specificities from the known mammalian GalTs. To study these enzymes, we cloned the pigeon liver cDNAs encoding alpha4GalT and beta4GalT by expression cloning and characterized these enzymes using the recombinant proteins. The deduced amino acid sequence of pigeon alpha4GalT has 58.2% identity to human alpha4GalT and 68.0 and 66.6% identity to putative alpha4GalTs from chicken and zebra finch, respectively. Unlike human and putative chicken alpha4GalTs, which possess globotriosylceramide synthase activity, pigeon alpha4GalT preferred to catalyze formation of the Gal alpha1-4Gal sequence on glycoproteins. In contrast, the sequence of pigeon beta4GalT revealed a type II transmembrane protein consisting of 438 amino acid residues, with no significant homology to the glycosyltransferases so far identified from mammals and chicken. However, hypothetical proteins from zebra finch (78.8% identity), frogs (58.9-60.4%), zebrafish (37.1-43.0%), and spotted green pufferfish (43.3%) were similar to pigeon beta4GalT, suggesting that the pigeon beta4GalT gene was inherited from the common ancestors of these vertebrates. The sequence analysis revealed that pigeon beta4GalT and its homologs form a new family of glycosyltransferases.
我们之前发现，鸽 IgG 具有独特的 N-糖链结构，其非还原末端含有 Gal alpha1-4Gal beta1-4Gal beta1-4GlcNAc 序列。该序列很可能是由假定的 alpha1,4-和 beta1,4-半乳糖基转移酶（GalTs）产生的，它们分别负责 N-糖链上 Gal alpha1-4Gal 和 Gal beta1-4Gal 序列的生物合成。由于在哺乳动物糖蛋白中未发现这种聚糖结构，因此产生这些聚糖的生物合成酶可能具有与已知的哺乳动物 GalTs 不同的底物特异性。为了研究这些酶，我们通过表达克隆克隆了鸽子肝脏 cDNA 编码 alpha4GalT 和 beta4GalT，并使用重组蛋白对这些酶进行了表征。鸽子 alpha4GalT 的推导氨基酸序列与人 alpha4GalT 具有 58.2%的同一性，与鸡和斑胸草雀的假定 alpha4GalTs 分别具有 68.0%和 66.6%的同一性。与具有神经节苷脂合酶活性的人 alpha4GalT 和假定的鸡 alpha4GalT 不同，鸽子 alpha4GalT 更倾向于催化糖蛋白上 Gal alpha1-4Gal 序列的形成。相比之下，鸽子 beta4GalT 的序列揭示了一种由 438 个氨基酸残基组成的 II 型跨膜蛋白，与迄今为止从哺乳动物和鸡中鉴定出的糖基转移酶没有显著同源性。然而，斑胸草雀（78.8%同一性）、青蛙（58.9-60.4%）、斑马鱼（37.1-43.0%）和斑点绿河豚（43.3%）的假设蛋白与鸽子 beta4GalT 相似，表明鸽子 beta4GalT 基因是从这些脊椎动物的共同祖先遗传下来的。序列分析表明，鸽子 beta4GalT 及其同源物形成了一个新的糖基转移酶家族。
