Another building block in the plant cell wall: Barley xyloglucan xyloglucosyl transferases link covalently xyloglucan and anionic oligosaccharides derived from pectin

Abstract

We report on the homo-and hetero-transglycosylation activities of the HvXET3 and HvXET4 xyloglucan xyloglucosyl transferases (XET; EC 2.4.1.207) from barley (Hordeum vulgare L.), and the visualisation of these activities in young barley roots using Alexa Fluor 488-labelled oligosaccharides. We discover that these isozymes catalyse the transglycosylation reactions with the chemically defined donor and acceptor substrates, specifically with the xyloglucan donor and the penta-galacturonide [a(1-4)GalAp] 5 acceptorthe homogalacturonan (pectin) fragment. This activity is supported by 3D molecular models of HvXET3 and HvXET4 with the docked XXXG donor and [a(1-4)GalAp] 5 acceptor substrates at the À4 to +5 subsites in the active sites. Comparative sequence analyses of barley isoforms and seed-localised TmXET6.3 from nasturtium (Tropaeolum majus L.) permitted the engineering of mutants of TmXET6.3 that could catalyse the hetero-transglycosylation reaction with the xyloglucan/[a(1-4)GalAp] 5 substrate pair, while wild-type TmXET6.3 lacked this activity. Expression data obtained by real-time quantitative polymerase chain reaction of HvXET transcripts and a clustered heatmap of expression profiles of the gene family revealed that HvXET3 and HvXET6 co-expressed but did not share the monophyletic origin. Conversely, HvXET3 and HvXET4 shared this relationship, when we examined the evolutionary history of 419 glycoside hydrolase 16 family members, spanning monocots, eudicots and a basal Angiosperm. The discovered hetero-transglycosylation activity in HvXET3 and HvXET4 with the xyloglucan/[a(1-4)GalAp] 5 substrate pair is discussed against the background of roles of xyloglucan-pectin heteropolymers and how they may participate in spatial patterns of cell wall formation and remodelling , and affect the structural features of walls.

Recent Advances in Understanding the Roles of Pectin as an Active Participant in Plant Signaling Networks

“…The presence of this xyloglucan–pectin linkage in most angiosperms, despite extensive changes in xyloglucan structure and differences in cell wall composition, suggests that this structure is evolutionarily conserved and may be required for effective cell wall structure and function. Enzymes that form xyloglucan–pectin bonds were also identified [ 68 ]. The molecular mass and composition of xyloglucans can be altered following their deposition into plant cell walls through the activities of xyloglucan endotransglycosylases (XETs) and xyloglucan endohydrolases (XEHs), forming the xyloglucan endotransglucosylase/hydrolase (XTH) group of enzymes [ 69 ].…”

“…The molecular mass and composition of xyloglucans can be altered following their deposition into plant cell walls through the activities of xyloglucan endotransglycosylases (XETs) and xyloglucan endohydrolases (XEHs), forming the xyloglucan endotransglucosylase/hydrolase (XTH) group of enzymes [ 69 ]. In barley, HvXET3 and HvXET4 have hetero-transglycosylation activity as well as homo-transglycosylation activity [ 68 ]. They use xyloglucans as donors and pectin fragments as acceptors to form a xyloglucan–pectin bond.…”

Primary Cell Wall Modifying Proteins Regulate Wall Mechanics to Steer Plant Morphogenesis

“…Thus, newly synthesized polymers are integrated into the wall without destabilizing it ( Chebli and Geitmann, 2017 ; Chebli et al, 2021 ). Although exactly how this integration is accomplished is unclear, it was proposed that self-assembly ( Cannon et al, 2008 ) and enzyme-mediated wall assembly ( Holland et al, 2020 ; Stratilova et al, 2020 ) play important roles in the integration process of cell walls. With regard to the self-assembly of cell walls, readers could find details in this review ( Murugesan et al, 2015 ).…”

Conservation of endo-glucanase 16 (EG16) activity across highly divergent plant lineages

“…Individual XETs and XEHs have been implicated in diverse morphological processes in plants, such as cell growth and expansion, fruit ripening, flower development, and seed germination (2,(4)(5)(6)(7)(8)(9)(10)(11). Interestingly, some homologs have been shown to catalyze heterotransglycosylation between xyloglucan and other saccharides (11)(12)(13)(14).…”