Defining natural factors that stimulate and inhibit cellulose: Xyloglucan hetero-transglucosylation

Klaus Herburger, Lenka Frankova, Martina Picmanova, Anzhou Xin, Frank Meulewaeter, Andrew D. Hudson, Stephen C. Fry

Research output: Contribution to journalArticlepeer-review

Abstract

Certain transglucanases can covalently graft cellulose and mixed-linkage -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 heterotransglucosylation, we studied Equisetum fluviatile hetero-trans-beta-glucanase (EfHTG), which exhibits both CXE and MXE activity, exceeding its XET (xyloglucan: xyloglucan homo-transglucosylation) 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~300% increased by addition of methanol-boiled Equisetum extracts; there was no similar effect when the same enzyme acted on soluble donors (MLG orxyloglucan). The methanol-stable factor is proposed to be expansin-like, a suggestion supported by observations of pH dependence. Screening numerou slow-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.
Original languageEnglish
Number of pages17
JournalThe Plant Journal
Early online date13 Dec 2020
DOIs
Publication statusE-pub ahead of print - 13 Dec 2020

Keywords

  • cell wall
  • cellulose
  • cellobiose
  • equisetum fluviatile
  • expansins
  • hemicelluloses
  • hetero-trans-β-glucanase
  • transglycosylation
  • mixed-linkage β-glucan
  • xyloglucan

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