A general liquid chromatography/mass spectroscopy-based assay for detection and quantitation of methyltransferase activity

Methyltransferases form a large class of enzymes, most of which use S-adenosylmethionine as the methyl donor. In fact, S-adenosylmethionine is second only to ATP in the variety of reactions for which it serves as a cofactor. Several methods to measure methyltransferase activity have been described,...

Full description

Saved in:
Bibliographic Details
Published in:Analytical biochemistry 2006-02, Vol.349 (1), p.112-117
Main Authors: Salyan, Mary Ellen K., Pedicord, Donna L., Bergeron, Laurie, Mintier, Gabe A., Hunihan, Lisa, Kuit, Kathy, Balanda, Lynn A., Robertson, Barbara J., Feder, John N., Westphal, Ryan, Shipkova, Petia A., Blat, Yuval
Format: Article
Language:English
Subjects:
Amino Acid Sequence
Catechol O-Methyltransferase - chemistry
Catechol O-Methyltransferase - metabolism
Catechol O-Methyltransferase - physiology
Chromatography, Liquid
COMT
Enzyme Activation
Humans
Kinetics
LC/MS
Methyltransferase
Molecular Sequence Data
S-adenosylhomocysteine
S-Adenosylhomocysteine - chemistry
S-Adenosylhomocysteine - metabolism
S-adenosylmethionine
Tandem Mass Spectrometry
Citations: Items that this one cites
Items that cite this one
Online Access:Get full text
Tags: Add Tag
No Tags, Be the first to tag this record!
Description
Summary:Methyltransferases form a large class of enzymes, most of which use S-adenosylmethionine as the methyl donor. In fact, S-adenosylmethionine is second only to ATP in the variety of reactions for which it serves as a cofactor. Several methods to measure methyltransferase activity have been described, most of which are applicable only to specific enzymes and/or substrates. In this work we describe a sensitive liquid chromatography/mass spectroscopy-based methyltransferase assay. The assay monitors the conversion of S-adenosylmethionine to S-adenosylhomocysteine and can be applied to any methyltransferase and substrate of interest. We used the well-characterized enzyme catechol O-methyltransferase to demonstrate that the assay can monitor activity with a variety of substrates, can identify new substrates, and can be used even with crude preparation of enzyme. Furthermore, we demonstrate the utility of the assay for kinetic characterization of enzymatic activity.
ISSN:0003-2697
1096-0309
DOI:10.1016/j.ab.2005.10.040