Xyloglucan endotransglycosylase: evidence for the existence of a relatively stable glycosyl-enzyme intermediate.
Xyloglucan endotransglycosylases (XETs) catalyse the breakdown of xyloglucan molecules predominantly by transglycosylation. In this process, fragments of cleaved polysaccharide are preferentially transferred to other xyloglucan molecules or their oligosaccharide subunits, with overall retention of the anomeric configuration of the glycosidic bond. In accordance with the theory, we propose that the cleavage and re-formation of the glycosidic bond in xyloglucan involves the formation of a glycosyl-enzyme intermediate which decomposes by transfer of the glycosyl moiety to a suitable carbohydrate acceptor. XETs from nasturtium seed cotyledons, mung bean hypocotyls and cauliflower florets interacted with xyloglucan to form complexes of high Mr as judged by gel-permeation chromatography. The nasturtium enzyme also showed evidence of XET-xyloglucan complex-formation according to anion-exchange chromatography and adsorption of the complex to filter paper on the basis of affinity of its xyloglucan moiety for cellulose. The XET-xyloglucan complex was stable in water, 6 M urea and acidic and alkaline buffers (pH 2.5-9.5), but readily decomposed by transferring its glycosyl moiety to xyloglucan-derived oligosaccharides or by incubation with the strong nucleophile imidazole at pH 3.8-9.6. These results strongly support the assumption that XET forms a relatively stable covalently linked glycosyl-enzyme intermediate.
Sulová
Z
Z
Institute of Chemistry, Slovak Academy of Sciences, 84238 Bratislava, Slovakia.
Takácová
M
M
Steele
N M
NM
Fry
S C
SC
Farkas
V
V
D002460:Cell Line; D002850:Chromatography, Gel; D002852:Chromatography, Ion Exchange; D007887:Fabaceae / Q000201:enzymology; D006031:Glycosylation; D016695:Glycosyltransferases / Q000737:chemistry* / Q000302:isolation & purification / Q000378:metabolism*; D007700:Kinetics; D008956:Models, Chemical; D010944:Plants / Q000201:enzymology*; D010946:Plants, Medicinal; D011485:Protein Binding; D012639:Seeds / Q000201:enzymology; D018724:Spinacia oleracea
D016695:Glycosyltransferases; C073693:xyloglucan endotransglycosylase