孤儿糖基转移酶的体外测定及其在鉴定Notch木糖基转移酶中的应用。

In vitro assays of orphan glycosyltransferases and their application to identify Notch xylosyltransferases.

作者信息

Sethi Maya K, Buettner Falk F R, Ashikov Angel, Bakker Hans

机构信息

Department of Cellular Chemistry, Hannover Medical School, Hannover, Germany.

出版信息

Methods Mol Biol. 2013;1022:307-20. doi: 10.1007/978-1-62703-465-4_23.

DOI:10.1007/978-1-62703-465-4_23

PMID:23765671

Abstract

Here we describe a systematic approach to determine the activity of putative glycosyltransferases with a focus on orphan members of the glycosyltransferase 8 family. An assay that measures the hydrolysis activity of glycoslytransferases can indicate the donor nucleotide sugar specificity without previous knowledge about the acceptor. Knowing the donor specificity, the acceptor specificity can subsequently be determined using synthetic acceptors. Three putative glycosyltransferases, now renamed GXYLT1, GXYLT2, and XXYLT1, have been identified this way as xylosyltransferases and in addition have been shown to act on O-glucosylated EGF repeats of Notch.

摘要

在此，我们描述了一种系统方法，用于确定推定的糖基转移酶的活性，重点关注糖基转移酶8家族的孤儿成员。一种测量糖基转移酶水解活性的测定方法可以在无需事先了解受体的情况下指示供体核苷酸糖的特异性。在了解供体特异性后，随后可使用合成受体来确定受体特异性。通过这种方式，已鉴定出三种推定的糖基转移酶，现重新命名为GXYLT1、GXYLT2和XXYLT1，它们是木糖基转移酶，此外还显示可作用于Notch的O-糖基化EGF重复序列。