Direct detection of rutin-degrading isozymes with polyacrylamide gel electrophoresis.
Rutin-degrading enzymes (RDEs) specifically hydrolyze the glycosidic linkages of rutin, producing quercetin and rutinose. Here we report a reliable and sensitive polyacrylamide gel electrophoresis and staining method for the detection of RDE isozymes, which is based on the aqueous solubility difference between rutin and quercetin, as well as the ultraviolet absorbance of quercetin. With this novel method, we achieved a detection limit of 12 ng with 107 U of RDE activity, enabling us to detect at least five RDE isozymes in tartary buckwheat seeds.
College of Life Sciences, Northwest A&F University, Yangling, Shaanxi 712100, China.
Yuping
Y
Li
Dandan
D
Deng
Xuebin
X
Zhang
Haina
H
Zhang
Cong
C
Wang
Peng
P
Chen
D015153:Blotting, Western; D004591:Electrophoresis, Polyacrylamide Gel / Q000379:methods*; D057075:Enzyme Assays / Q000379:methods*; D019612:Fagopyrum / Q000201:enzymology* / Q000378:metabolism; D007527:Isoenzymes / Q000302:isolation & purification / Q000378:metabolism; D057230:Limit of Detection; D012431:Rutin / Q000302:isolation & purification / Q000378:metabolism*
D007527:Isoenzymes; D012431:Rutin
Isozyme; PAGE; Rutin-degrading enzyme; Western blotting