Defining natural factors that stimulate and inhibit cellulose:xyloglucan hetero‐transglucosylation

Abstract

Certain transglucanases can covalently graft cellulose and mixed-linkage b-glucan (MLG) as donor substrates onto xyloglucan as acceptor substrate and thus exhibit cellulose:xyloglucan endotransglucosylase (CXE) and MLG:xyloglucan endotransglucosylase (MXE) activities in vivo and in vitro. However, missing information on factors that stimulate or inhibit these hetero-transglucosylation reactions limits our insight into their biological functions. To explore factors that influence hetero-transglucosylation, we studied Equisetum fluviatile hetero-trans-b-glucanase (EfHTG), which exhibits both CXE and MXE activity, exceeding its xyloglucan:xyloglucan homo-transglucosylation (XET) activity. Enzyme assays employed radiolabelled and fluorescently labelled oligomeric acceptor substrates, and were conducted in vitro and in cell walls (in situ). With whole denatured Equisetum cell walls as donor substrate, exogenous EfHTG (extracted from Equisetum or produced in Pichia) exhibited all three activities (CXE, MXE, XET) in competition with each other. Acting on pure cellulose as donor substrate, the CXE action of Pichia-produced EfHTG was up to approximately 300% increased by addition of methanol-boiled Equisetum extracts; there was no similar effect when the same enzyme acted on soluble donors (MLG or xyloglucan). The methanol-stable factor is proposed to be expansin-like, a suggestion supported by observations of pH dependence. Screening numerous low-molecular-weight compounds for hetero-transglucanase inhibition showed that cellobiose was highly effective, inhibiting the abundant endogenous CXE and MXE (but not XET) action in Equisetum internodes. Furthermore, cellobiose retarded Equisetum stem elongation, potentially owing to its effect on hetero-transglucosylation reactions. This work provides insight and tools to further study the role of cellulose hetero-transglucosylation in planta by identifying factors that govern this reaction.

Forgotten Actors: Glycoside Hydrolases During Elongation Growth of Maize Primary Root

“…Maize genes are given in black font for expressed genes and gray for those having TGR values lower than 16 in all the analyzed root samples, and Arabidopsis thaliana genes are given in red. Characterized XTHs of barley ( Hrmova et al, 2007 , 2009 ), Equisetum ( Herburger et al, 2021 ), nasturtium ( Crombie et al, 1998 ), and Brachypodium ( Fan et al, 2018 ) are shown in purple, green, blue, and magenta, respectively. Numbers indicate ultrafast bootstrap support values for some branches.…”

“…Moreover, in Brachypodium , BdXTH8 accumulates in elongating tissues characterized by high amounts of MLG ( Fan et al, 2018 ). The Equisetum homolog of BdXTH8 (EfHTG) is able to use cellulose as the donor substrate and exhibits the dominance of cellulose:XyG endotransglucosylase activity over XyG:XyG homo-transglucosylation activity ( Herburger et al, 2021 ). HvXTH5 also encodes an XTH capable of using cellulose as the donor substrate although with a rate lower than that for XyG:XyG endotransglycosylation ( Hrmova et al, 2007 ).…”

Cutin:cutin-acid endo-transacylase (CCT), a cuticle-remodelling enzyme activity in the plant epidermis

“…Incorporation was maximal in the apoplastic pH range (4.5-6.5), and the reaction was strongly diminished if the epidermis was heat-denatured (Figure 3a; P < 0.05), compatible with the involvement of an apoplastic enzyme, such enzymes often being relatively heat-stable [57]. Such a pH optimum is close to that of the apoplastic CUS1 [58], but appreciably lower than that of the system reported by [56], which was dependent on ATP, CoA and a protoplasmic HFA-CoA synthetase.…”

Chara — a living sister to the land plants with pivotal enzymic toolkit for mannan and xylan remodelling

“…The results thus indicate that the endo-enzyme transβ-mannanase, despite having a lower affinity (higher K M ) for Man 6 , was ultimately more active than endo-enzyme trans-β-mannosidase. plant kingdom, though some are confined to (or only prominent in) certain taxa such as trans-β-mannanase in some ferns, hornworts and lycophytes MXE andCXE in Equisetum (Simmons et al, 2015;Herburger et al, 2021). In this work, we extended the search for such enzymes to a charophytic alga, Chara vulgaris.…”

“…Heterotransglycanases can be of great interest in that they interlink dissimilar polysaccharides within the cell wall — for instance bonding cellulose to xyloglucan and thus potentially anchoring xyloglucan to the cellulosic microfibrils (Herburger et al, 2021 ).…”