Department of Chemistry, University of Natural Resources and Life Sciences, 1190 Vienna, Austria.
Department of Biotechnology, University of Natural Resources and Life Sciences, 1190 Vienna, Austria.
Molecules. 2023 Jan 5;28(2):552. doi: 10.3390/molecules28020552.
UDP-Gal: glycoprotein-N-acetylgalactosamine β-1,3-galactosyltransferase (T-synthase, EC 2.4.1.122) catalyses the transfer of the monosaccharide galactose from UDP-Gal to GalNAc-Ser/Thr, synthesizing the core 1 mucin type O-glycan. Such glycans play important biological roles in a number of recognition processes. The crucial role of these glycans is acknowledged for mammals, but a lot remains unknown regarding invertebrate and especially mollusc O-glycosylation. Although core O-glycans have been found in snails, no core 1 β-1,3-galactosyltransferase has been described so far. Here, the sequence of the enzyme was identified by a BlastP search of the NCBI database using the human T-synthase sequence (NP_064541.1) as a template. The obtained gene codes for a 388 amino acids long transmembrane protein with two putative N-glycosylation sites. The coding sequence was synthesised and expressed in Sf9 cells. The expression product of the putative enzyme displayed core 1 β-1,3-galactosyltransferase activity using pNP-α-GalNAc as the substrate. The enzyme showed some sequence homology (49.40% with , 53.69% with and 49.14% with ) and similar biochemical parameters with previously characterized T-synthases from other phyla. In this study we present the identification, expression and characterisation of the UDP-Gal: glycoprotein-N-acetylgalactosamine β-1,3-galactosyltransferase from the fresh-water snail , which is the first cloned T-synthase from mollusc origin.
UDP-Gal: 糖蛋白-N-乙酰半乳糖胺 β-1,3-半乳糖基转移酶（T-合成酶，EC 2.4.1.122）催化单糖半乳糖从 UDP-Gal 转移到 GalNAc-Ser/Thr，合成核心 1 粘蛋白型 O-聚糖。这些聚糖在许多识别过程中发挥着重要的生物学作用。这些聚糖对于哺乳动物的关键作用已得到认可，但对于无脊椎动物，特别是软体动物的 O-糖基化，仍有很多未知之处。尽管已经在蜗牛中发现了核心 O-聚糖，但迄今为止尚未描述核心 1 β-1,3-半乳糖基转移酶。在这里，使用人类 T-合成酶序列（NP_064541.1）作为模板，通过对 NCBI 数据库进行 BlastP 搜索，鉴定了该酶的序列。获得的基因编码一个 388 个氨基酸长的跨膜蛋白，带有两个假定的 N-糖基化位点。合成并在 Sf9 细胞中表达了编码序列。假定酶的表达产物使用 pNP-α-GalNAc 作为底物显示出核心 1 β-1,3-半乳糖基转移酶活性。该酶与其他门的已表征 T-合成酶具有一定的序列同源性（与 49.40%，与 53.69%，与 49.14%），并且具有相似的生化参数。在这项研究中，我们介绍了来自淡水蜗牛 的 UDP-Gal: 糖蛋白-N-乙酰半乳糖胺 β-1,3-半乳糖基转移酶的鉴定、表达和特性，这是第一个从软体动物来源克隆的 T-合成酶。
