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Title: Transglucosylation of cell wall polysaccharides in equisetum fluviatile
Author: Mohler, Kyle Edward
Awarding Body: University of Edinburgh
Current Institution: University of Edinburgh
Date of Award: 2012
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Plant cell walls determine cellular shape and provide structural support for the entire plant. Polysaccharides, comprising the major components of the wall, are actively remodelled throughout development. Xyloglucan endotransglucosylase (XET)/hydrolase (XTH, EC cleaves xyloglucan (XyG), the donor substrate, and attaches a portion to another XyG chain, the acceptor substrate. Recently, a novel transglucosylase called mixed-linkage β-glucan (MLG) : XyG endotransglucosylase (MXE) was discovered in horsetails (Equisetum spp.) that could attach a portion of MLG to XyG, resulting in a hetero-polymer product. My aims were to further investigate the nature of this activity, biochemically characterize the enzyme, and explore its physiological role. MXE activity was attributable to an enzyme unlike Equisetum XTHs. MXE had a p1 of 4.1 (XTHs were 6.6-9), a pH optimum of 6.3 (XTHs preferred 5.5), and had higher activity using smaller oligosaccharide acceptor substrates like XXXGol (XTHs were more active using XLLGol). Importantly, the MXE protein was shown to utilize both MLG and XyG as donor substrates, and therefore have both MXE and XET activity. Also, the enzyme was capable of using various glucan oligosaccharides (O) as substrates, including MLGO, XyGO, and cello-O, but not laminari-O. By using a novel ex vivo approach, the proportion of extractable MXE product to XET product was found to increase in older tissues. Transglucosylase products were localized in sclerenchyma and structural parenchyma by in situ assays, implying a strenghening function for MXE. Surprisingly, another novel activity was discovered that could covalently attach cellulose to XyG, and termed cellulose : xyloglucan endotransglucosylase (CXE). This activity was attributed to the MXE enzyme, implying that the protein is a promiscuous endotransglucosylase. The presence of CXE in other plants has not yet been tested. Besides being a novel discovery in plant cell biology, the modification of cellulose has applications in a number of industries.
Supervisor: Fry, Stephen; Hudson, Andrew Sponsor: Darwin Trust of Edinburgh
Qualification Name: Thesis (Ph.D.) Qualification Level: Doctoral
EThOS ID:  DOI: Not available
Keywords: plant cell wall ; transglucosylation ; xyloglucan ; mixed-linkage glucan ; equisetum