Open AccessSynthesis of a New Photoaffinity Probe, 5‐Azido‐[ 32 P]UDPxylose, by UDPglucuronate Carboxylyase from Wheat Germ
Author(s) -
Kyossev Zhetcho N.,
Drake Richard R.,
Kyosseva Svetlana V.,
Elbein Alan D.
Publication year - 1995
Publication title -
european journal of biochemistry
Language(s) - English
Resource type - Journals
eISSN - 1432-1033
pISSN - 0014-2956
DOI - 10.1111/j.1432-1033.1995.0109o.x
Subject(s) - chemistry , wheat germ , yield (engineering) , photoaffinity labeling , ion chromatography , enzyme , chromatography , specific activity , biochemistry , binding site , materials science , metallurgy
The enzyme, UDPglucuronic acid carboxylyase (EC 4.1.1.35), was extensively purified from wheat germ, and was used to convert 5‐azido‐[ 32 P]UDPglucuronic acid to 5‐azido‐[ 32 P]UDPxylose, for use as a new photoaffinity probe. The carboxylyase was purified approximately 1200‐fold using conventional methods, and the enzyme preparation, at the final stage of purification, was stable to storage at −20°C for at least 9 months with little or no loss of activity. The partially purified carboxylyase catalyzed the conversion of 5‐azido‐[ 32 P]UDPglucuronic acid to 5‐azido‐[ 32 P]UDPxylose in good yield, and the UDPxylose probe was purified by ion‐exchange chromatography, and characterized. The newly synthesized photoaffinity analog, 5‐azido‐[ 32 P]UDPxylose, should be a valuable tool in the purification of various xylosyltransferases.