Research & Faculty

Default Header Image

Synthesis and evaluation of a series of 6-chloro-4-methylumbelliferyl glycosides as fluorogenic reagents for screening metagenomic libraries for glycosidase activity

TitleSynthesis and evaluation of a series of 6-chloro-4-methylumbelliferyl glycosides as fluorogenic reagents for screening metagenomic libraries for glycosidase activity
Publication TypeJournal Article
Year of Publication2016
AuthorsChen, H-M, Armstrong, Z, Hallam, SJ, Withers, SG
JournalCARBOHYDRATE RESEARCH
Volume421
Pagination33-39
Date PublishedFEB 8
ISSN0008-6215
Abstract

Screening of large enzyme libraries such as those derived from metagenomic sources requires sensitive substrates. Fluorogenic glycosides typically offer the best sensitivity but typically must be used in a stopped format to generate good signal. Use of fluorescent phenols of pKa < 7, such as halogenated coumarins, allows direct screening at neutral pH. The synthesis and characterisation of a set of nine different glycosides of 6-chloro-4-methylumbelliferone are described. The use of these substrates in a pooled format for screening of expressed metagenomic libraries yielded a ``hit rate{''} of 1 in 60. Hits were then readily deconvoluted with the individual substrates in a single plate to identify specific activities within each clone. The use of such a collection of substrates greatly accelerates the screening process. (C) 2015 Elsevier Ltd. All rights reserved.

DOI10.1016/j.carres.2015.12.010