Visualization of the activity of xyloglucan endotransglycosylase (XET) isoenzymes after gel electrophoresis

A protocol is described which allows the detection of transglycosylase isoe nzyme activity, Crude extracts of cauliflower (Brassica oleracea var. botry tis) fiorets were treated with 1 M sodium chloride, filtered through cellul ose acetate, dialysed, concentrated and subjected to isoelectric focusing u nder non-denaturing conditions in a polyacrylamide gel. After electrophores is, the gel was overlaid with a test paper impregnated with a mixture of hi gh-M-r xyloglucan and a sulphorhodamine-conjugated oligosaccharide of xylog lucan. Transglycosylation caused the production of sulphorhodamine-labelled polysaccharide, which remained hound to the test paper after prolonged was hing in water, revealing eight fluorescent hands corresponding to XET isoen zymes with pi values of 8.8, 8.6, 7.8, 7.6, 7.5, 7.1, 6,7 and 6.6. Copyrigh t (C) 1999 John Wiley & Sons, Ltd.