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Anal Biochem. 2004 Sep 1;332(1):109-15.

Enzyme-coupled assay of acetylxylan esterases on monoacetylated 4-nitrophenyl beta-D-xylopyranosides.

Author information

1
Institute of Chemistry, Slovak Academy of Sciences, Dúbravská cesta 9, 84538 Bratislava, Slovakia. chempbsa@savba.sk

Abstract

Three different monoacetates of 4-nitrophenyl beta-D-xylopyranoside were tested as substrates for beta-xylosidase and for microbial carbohydrate esterases and a series of non-hemicellulolytic esterases. The acetyl group in 2-O-acetyl, 3-O-acetyl, and 4-O-acetyl 4-nitrophenyl beta-D-xylopyranoside makes the glycoside resistant to the action of beta-xylosidase (EC 3.2.1.37). This fact was explored to introduce a new enzyme-coupled assay of acetylxylan esterases (EC 3.1.1.72) and other carbohydrate-deacetylating enzymes. The deacetylation converts the monoacetates into the substrate of beta-xylosidase, the auxiliary enzyme. The effect of the acetyl group migration along the xylopyranoid ring in aqueous media can be avoided by shortening the assay duration. The assay enables an easy examination of the positional specificity of the enzymes, which is important for classification of acetylxylan esterases and for elucidation of the structure-function relationship among carbohydrate esterases in general. Non-hemicellulolytic esterases showed different positional specificity of deacetylation than did acetylxylan esterases.

PMID:
15301955
DOI:
10.1016/j.ab.2004.04.022
[Indexed for MEDLINE]

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