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

✍ Scribed by Pietro P. M. Iannetta; Stephen C. Fry

Publisher: John Wiley and Sons
Year: 1999
Tongue: English
Weight: 69 KB
Volume: 10
Category: Article
ISSN: 0958-0344
DOI: 10.1002/(sici)1099-1565(199909/10)10:5<238::aid-pca459>3.0.co;2-x

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✦ Synopsis

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.

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