TY - JOUR
T1 - Visualization of the activity of xyloglucan endotransglycosylase (XET) isoenzymes after gel electrophoresis
AU - Iannetta, Pietro P.M.
AU - Fry, Stephen C.
PY - 1999
Y1 - 1999
N2 - A protocol is described which allows the detection of transglycosylase isoenzyme activity. Crude extracts of cauliflower (Brassica oleracea var. botrytis) florets were treated with 1 M sodium chloride, filtered through cellulose acetate, dialysed, concentrated and subjected to isoelectric focusing under non-denaturing conditions in a polyacrylamide gel. After electrophoresis, the gel was overlaid with a test paper impregnated with a mixture of high-M(r) xyloglucan and a sulphorhodamine-conjugated oligosaccharide of xyloglucan. Transglycosylation caused the production of sulphorhodamine-labelled polysaccharide, which remained bound to the test paper after prolonged washing in water, revealing eight fluorescent bands corresponding to XET isoenzymes with pI values of 8.8, 8.6, 7.8, 7.6, 7.3, 7.1, 6.7 and 6.6.
AB - A protocol is described which allows the detection of transglycosylase isoenzyme activity. Crude extracts of cauliflower (Brassica oleracea var. botrytis) florets were treated with 1 M sodium chloride, filtered through cellulose acetate, dialysed, concentrated and subjected to isoelectric focusing under non-denaturing conditions in a polyacrylamide gel. After electrophoresis, the gel was overlaid with a test paper impregnated with a mixture of high-M(r) xyloglucan and a sulphorhodamine-conjugated oligosaccharide of xyloglucan. Transglycosylation caused the production of sulphorhodamine-labelled polysaccharide, which remained bound to the test paper after prolonged washing in water, revealing eight fluorescent bands corresponding to XET isoenzymes with pI values of 8.8, 8.6, 7.8, 7.6, 7.3, 7.1, 6.7 and 6.6.
KW - Hydrolase
KW - Isoelectric focusing
KW - Native polyacrylamide gel electrophoresis
KW - Transglycosylase
KW - Xyloglucan
KW - Zymogram
UR - http://www.scopus.com/inward/record.url?scp=0032878779&partnerID=8YFLogxK
U2 - 10.1002/(SICI)1099-1565(199909/10)10:5<238::AID-PCA459>3.0.CO;2-X
DO - 10.1002/(SICI)1099-1565(199909/10)10:5<238::AID-PCA459>3.0.CO;2-X
M3 - Article
AN - SCOPUS:0032878779
SN - 0958-0344
VL - 10
SP - 238
EP - 240
JO - Phytochemical Analysis
JF - Phytochemical Analysis
IS - 5
ER -