Award details

Cross-linking of extracellular feruloyl-polysaccharides in maize by oxidative coupling and the formation of dehydrodiferulate-sugar ether bonds

Reference	BB/C505791/1
Principal Investigator / Supervisor	Professor Stephen Fry
Co-Investigators / Co-Supervisors
Institution	University of Edinburgh
Department	Inst for Molecular Plant Science
Funding type	Research
Value (£)	253,630
Status	Completed
Type	Research Grant
Start date	01/03/2005
End date	29/02/2008
Duration	36 months

Abstract

The cell walls of agriculturally and nutritionally valuable cereals and grasses are characteristic structural elements of gramineous tissue that govern its texture. The majority of past cell wall studies have dealt with dicotyledons. However, Gramineous primary cell walls (PCWs) differ chemically in several fundamental ways from those of dicots: for example, the former uniquely have mixed-linkage beta-glucans; they have more xylans, ferulate and beta-expansin; and they have less pectin, xyloglucan, extensin and alpha-expansin. It is therefore timely, and commercially appropriate, to focus some additional basic research on PCWs of the Gramineae, ie. major world crops. The project deals with the cross-linking of non-cellulosic polysaccharides (especially xylans) of the gramineous PCW via feruloyl ester side-chains. Such cross-linking is thought to be centrally involved in several important processes including: controlling cell expansion and thus plant growth; restricting susceptibility to penetration by pathogens and thus improving disease-resistance; and promoting cell-cell adhesion and thus consolidating tissues. Studying the mechanisms and control of the formation of these cross-links is complicated by the fact that PCW components are insoluble both before and after cross-linking. To facilitate the demonstration of true (inter-polymeric) cross-linking, we will exploit a useful model system recently developed in our lab ¿ the cross-linking of soluble extracellular polysaccharides (SEPs) in the medium of cultured maize cells. Although SEPs have often been proposed as models of PCW polysaccharide structures, little use has been made of their in-vivo reactions to model wall metabolism. Maize SEPs undergo sudden cross-linking, in a highly controlled fashion, after several days in the culture. Although recent evidence indicates that the cross-linking is dependent on oxidative phenolic coupling, which would produce dehydrodiferuloyl esters, the cross-linked SEPs cannot bereturned to their original molecular weight by de-esterification. Instead, restoring their original molecular weight requires more severe alkali, similar to that necessary for splitting p-hydroxybenzyl-sugar ether bonds. This indicates the presence of ether-like cross-links (probably in addition to ester cross-links). We now aim to describe the chemical nature of these cross-linking processes, especially the formation of ether-like bonds between dehydrodiferulates and polysaccharides. The ester and ether bonds formed will be traced by in-vivo radiolabelling using exogenous [14C]cinnamate and [3H]arabinose. They will be characterised radiochemically after graded enzymic, acid and alkaline hydrolysis, and if possible isolated in quantities sufficient for MS and NMR studies. A major question to be addressed is whether the formation of phenol-sugar ether bonds is initiated, or at least steered, by novel proteins. If either the reaction of a QM (an esterified, quinone-methide-type, dehydrodimer or ¿trimer of ferulate) with a sugar residue to form an ether bond is enzyme-paralysed, as we suggest, such an enzyme would be a novel carbon-oxygen lyase (acting in the synthase direction). We will carry out assays designed to detect such an enzyme activity. A second possibility is the existence of a dirigent-like protein. An ability to assay the lyase or dirigent would open the way to the future identification of their genes, and thus our future ability to modify their action genetically or biotechnologically. We will also explore the control of ferulate-based ester and ether cross-linking, determining whether any or all H202, peroxidase or the concentration of other polysaccharides limits the cross-linking. Although the project is not targeted at any one specific gramineous crop, the potential outcome of the work is an ability to improve the food/fodder quality of cereals and grasses by modification of their structural (cell wall) components.

Summary

unavailable

Committee	Closed Committee - Agri-food (AF)
Research Topics	X – not assigned to a current Research Topic
Research Priority	X – Research Priority information not available
Research Initiative	X - not in an Initiative
Funding Scheme	X – not Funded via a specific Funding Scheme

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