A rationally engineered yeast pyruvyltransferase Pvg1p introduces sialylation-like properties in neo-human-type complex oligosaccharide.
A rationally engineered yeast pyruvyltransferase Pvg1p introduces sialylation-like properties in neo-human-type complex oligosaccharide.
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DOI：
10.1038/srep26349
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发表时间：
2016-05-19
影响因子：
4.6
通讯作者：
Takegawa K
中科院分区：
文献类型：
--
作者：
Higuchi Y;Yoshinaga S;Yoritsune K;Tateno H;Hirabayashi J;Nakakita S;Kanekiyo M;Kakuta Y;Takegawa K
从原核生物到真核生物，广泛存在于寡糖末端的丙酮酰化，在细胞表面赋予负电荷，并且似乎在功能上类似于唾液酸化，唾液酸化存在于人型复合寡糖的末端。然而，详细的分子机制，潜在的乙酰基化还没有得到很好的澄清。在这里，我们首先以2.46 μ m的分辨率确定了裂殖酵母酰基转移酶Pvg 1 p的晶体结构。随后，通过结合分子模拟与活性位点残基的突变分析，我们获得了一个Pvg 1 p突变体（Pvg 1 pH 168 C），有效地转移到一个人型复杂的糖肽的乙酰基部分。所得的唾液酸化的人型复合糖肽识别凝集素阵列上的类似凝集素作为α 2，6-唾液酸糖肽。这种新产生的人型复合寡糖的乙酰化将为糖生物工程提供新的方法。
Pyruvylation onto the terminus of oligosaccharide, widely seen from prokaryote to eukaryote, confers negative charges on the cell surface and seems to be functionally similar to sialylation, which is found at the end of human-type complex oligosaccharide. However, detailed molecular mechanisms underlying pyruvylation have not been clarified well. Here, we first determined the crystal structure of fission yeast pyruvyltransferase Pvg1p at a resolution of 2.46 Å. Subsequently, by combining molecular modeling with mutational analysis of active site residues, we obtained a Pvg1p mutant (Pvg1pH168C) that efficiently transferred pyruvyl moiety onto a human-type complex glycopeptide. The resultant pyruvylated human-type complex glycopeptide recognized similar lectins on lectin arrays as the α2,6-sialyl glycopeptides. This newly-generated pyruvylation of human-type complex oligosaccharides would provide a novel method for glyco-bioengineering.