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Methods Mol Biol. 2017;1588:3-14. doi: 10.1007/978-1-4939-6899-2_1.

A Low-Volume, Parallel Copper-Bicinchoninic Acid (BCA) Assay for Glycoside Hydrolases.

Author information

1
Michael Smith Laboratories, University of British Columbia, 2185 East Mall, Vancouver, BC, V6T 1Z4, Canada.
2
Department of Chemistry, University of British Columbia, 2185 East Mall, Vancouver, BC, Canada, V6T 1Z4.
3
Michael Smith Laboratories, University of British Columbia, 2185 East Mall, Vancouver, BC, V6T 1Z4, Canada. brumer@msl.ubc.ca.
4
Department of Chemistry, University of British Columbia, 2185 East Mall, Vancouver, BC, Canada, V6T 1Z4. brumer@msl.ubc.ca.
5
Department of Biochemistry and Molecular Biology, University of British Columbia, 2185 East Mall, Vancouver, BC, Canada, V6T 1Z4. brumer@msl.ubc.ca.

Abstract

The quantitation of liberated reducing sugars by the copper-bicinchoninic acid (BCA) assay provides a highly sensitive method for the measurement of glycoside hydrolase (GH) activity, particularly on soluble polysaccharide substrates. Here, we describe a straightforward method adapted to low-volume polymerase chain reaction (PCR) tubes which enables the rapid, parallel determination of GH kinetics in applications ranging from initial activity screening and assay optimization, to precise Michaelis-Menten analysis.

KEYWORDS:

Carbohydrate-active enzymes (CAZymes); Copper-bicinchoninic acid (BCA); Enzymology; Glycosidase; Glycoside hydrolase (GH); Polysaccharide; Reducing sugar

PMID:
28417356
DOI:
10.1007/978-1-4939-6899-2_1
[Indexed for MEDLINE]

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