Department of Chemistry, University of British Columbia, Vancouver, British Columbia V6T 1Z3, Canada
Michael Smith Laboratories, University of British Columbia, Vancouver, British Columbia V6T 1Z4, Canada.
J Biol Chem. 2019 Nov 1;294(44):16400-16415. doi: 10.1074/jbc.RA119.010628. Epub 2019 Sep 17.
α-Linked GalNAc (α-GalNAc) is most notably found at the nonreducing terminus of the blood type-determining A-antigen and as the initial point of attachment to the peptide backbone in mucin-type -glycans. However, despite their ubiquity in saccharolytic microbe-rich environments such as the human gut, relatively few α--acetylgalactosaminidases are known. Here, to discover and characterize novel microbial enzymes that hydrolyze α-GalNAc, we screened small-insert libraries containing metagenomic DNA from the human gut microbiome. Using a simple fluorogenic glycoside substrate, we identified and characterized a glycoside hydrolase 109 (GH109) that is active on blood type A-antigen, along with a new subfamily of glycoside hydrolase 31 (GH31) that specifically cleaves the initial α-GalNAc from mucin-type -glycans. This represents a new activity in this GH family and a potentially useful new enzyme class for analysis or modification of -glycans on protein or cell surfaces.
α-连接的半乳糖胺（α-GalNAc）最显著的存在于血型决定 A 抗原的非还原末端，并且作为黏蛋白型聚糖中与肽主链连接的初始点。然而，尽管它们在富含糖解微生物的环境（如人类肠道）中无处不在，但已知的α-乙酰半乳糖胺酶相对较少。在这里，为了发现和表征水解α-GalNAc 的新型微生物酶，我们筛选了含有人类肠道微生物组宏基因组 DNA 的小插入文库。使用简单的荧光糖基化物底物，我们鉴定和表征了一种糖苷水解酶 109（GH109），它对血型 A 抗原具有活性，以及一种新的糖苷水解酶 31（GH31）亚家族，它特异性地从黏蛋白型聚糖中切割初始的α-GalNAc。这代表了该 GH 家族中的一种新活性，并且是一种潜在有用的新酶类，可用于分析或修饰蛋白质或细胞表面上的聚糖。
