Trans‐α‐xylosidase and trans‐β‐galactosidase activities, widespread in plants, modify and stabilize xyloglucan structures

Abstract

SUMMARYCell-wall components are hydrolysed by numerous plant glycosidase and glycanase activities. We investigated whether plant enzymes also modify xyloglucan structures by transglycosidase activities. Diverse angiosperm extracts exhibited transglycosidase activities that progressively transferred single sugar residues between xyloglucan heptasaccharide (XXXG or its reduced form, XXXGol) molecules, at 16 lM and above, creating octato decasaccharides plus smaller products. We measured remarkably high transglycosylation:hydrolysis ratios under optimized conditions. To identify the transferred monosaccharide(s), we devised a dual-labelling strategy in which a neutral radiolabelled oligosaccharide (donor substrate) reacted with an amino-labelled non-radioactive oligosaccharide (acceptor substrate), generating radioactive cationic products. For example, 37 lM [Xyl-3 H]XXXG plus 1 mM XXLG-NH 2 generated 3 H-labelled cations, demonstrating xylosyl transfer, which exceeded xylosyl hydrolysis 1.6-to 7.3-fold, implying the presence of enzymes that favour transglycosylation. The transferred xylose residues remained a-linked but were relatively resistant to hydrolysis by plant enzymes. Driselase digestion of the products released a trisaccharide (a-[ 3 H]xylosylisoprimeverose), indicating that a new xyloglucan repeat unit had been formed. In similar assays, [Gal-3 H]XXLG and [Gal-3 H]XLLG (but not [Fuc-3 H]XXFG) yielded radioactive cations. Thus plants exhibit trans-a-xylosidase and trans-b-galactosidase (but not trans-a-fucosidase) activities that graft sugar residues from one xyloglucan oligosaccharide to another. Reconstructing xyloglucan oligosaccharides in this way may alter oligosaccharin activities or increase their longevity in vivo. Trans-a-xylosidase activity also transferred xylose residues from xyloglucan oligosaccharides to long-chain hemicelluloses (xyloglucan, water-soluble cellulose acetate, mixed-linkage b-glucan, glucomannan and arabinoxylan). With xyloglucan as acceptor substrate, such an activity potentially affects the polysaccharide's suitability as a substrate for xyloglucan endotransglucosylase action and thereby modulates cell expansion. We conclude that certain proteins annotated as glycosidases can function as transglycosidases.

Boron bridging of rhamnogalacturonan‐II is promoted in vitro by cationic chaperones, including polyhistidine and wall glycoproteins

“…Given that no boron‐metabolizing enzyme is yet known, we undertook a preliminary study of the possibility that a plant protein might catalyse the boron‐dependent dimerization of RG‐II, thus ‘mimicking’ the action of Pb 2+ . The potential source was an extract of parsley shoots, a proven rich source of numerous cell‐wall‐directed enzyme activities including xyloglucan endotransglucosylase (XET), glycosidases, glycanases, transglycosidases, transglycanases and pectin methylesterase ( GHATA base, ; Franková & Fry, , , ; D. Chormova & S. C. Fry, unpublished). When the parsley extract was incubated for 0–24 h with [ 3 H]RG‐II (Fig.…”

Overexpression of the grapevine PGIP1 in tobacco results in compositional changes in the leaf arabinoxyloglucan network in the absence of fungal infection

“…Xyloglucan modifications in muro have been well studied in Arabidopsi s and mutants with changes in glycosidase activity, such as the Atxyl1, Atbgal10 and axy8 mutants [52-54] show significant alterations in the composition of the XEG-soluble polymers. It has recently been shown that trans-α-xylosidases and trans-β-galactosidases can act directly on the xyloglucan polysaccharide to cause modification [55]. …”

Glycosynthesis in a waterworld: new insight into the molecular basis of transglycosylation in retaining glycoside hydrolases

“…This catalytic sluggishness is likely to be correlated with more energy-demanding TS (for both glycosylation and deglycosylation steps), which lower overall catalytic turnover. mixed activity) [70][71][72][73][74][75]. Of course, this is a rather simplistic view of enzyme evolution and other data suggest that enzymes might have evolved in both directions [68,69], and indeed some GHs (mainly from plants) display both hydrolysis and transglycosylation activities and thus present intermediate cases (i.e.…”