Control of xyloglucan endotransglucosylase activity by salts and anionic polymers |
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Authors: | Takumi?Takeda,Stephen?C.?Fry author-information" > author-information__contact u-icon-before" > mailto:S.Fry@ed.ac.uk" title=" S.Fry@ed.ac.uk" itemprop=" email" data-track=" click" data-track-action=" Email author" data-track-label=" " >Email author |
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Affiliation: | (1) The Edinburgh Cell Wall Group, ICMB, The University of Edinburgh, Daniel Rutherford Building, The Kings Buildings, Edinburgh, EH9 3JH, UK;(2) Present address: Department of Biology, Pennsylvania State University, 208 Mueller Lab, University Park, PA 16802, USA |
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Abstract: | Crude extracts of cauliflower florets had high xyloglucan endotransglucosylase (XET) activity, but this was largely lost after partial purification and de-salting. Activity was restored (promoted up to 40-fold) by any of a wide variety of inorganic and organic salts. Optimum concentrations for Na+, K+ and NH4+ salts were typically ~300 mM. The chlorides of Ca2+, Mg2+, Al3+ and La3+ were optimally active at lower concentrations (e.g. 0.1 mM LaCl3), but became inhibitory at higher concentrations (e.g. 5 mM LaCl3). Some anionic polysaccharides at 0.04–0.2% w/v (e.g. gum arabic, pectin and hypochlorite-oxidised xyloglucan) promoted the XET activity of de-salted enzyme, especially if a sub-optimal concentration of NaCl was also present; others (e.g. homogalacturonan, 4-O-methyl-glucuronoxylan and alginate) were inhibitory. Similar ionic effects were noted on the XET activity of the Arabidopsis protein XTH24 (heterologously expressed by insect cells); in this case carboxymethylcellulose was also stimulatory. To look for endogenous modulators of XET activity, we prepared a cold-water extract of cauliflower florets; after boiling and centrifugation, the supernatant [boiled cauliflower preparation (BCP)] promoted the XET activity of de-salted cauliflower enzyme and of XTH24. About half the activator present in BCP was an ethanol-precipitable, anionic polymer of apparent Mr <5,000. After acid hydrolysis the polymer yielded much arabinose and galactose, and small amounts of galacturonic and glucuronic acids amino acids were also present. The polymer may thus contain arabinogalactan-proteins. We suggest that acidic polymers and/or other apoplastic ions are naturally occurring regulators of XET action in vivo, and may thus control cell wall assembly, loosening, and growth.Abbreviations AGP Arabinogalactan-protein - BCP Boiled cauliflower preparation (cold-water-extract of cauliflower florets that was then boiled) - CMC Carboxymethylcellulose - DE Degree of esterification - GalA Galacturonic acid - GlcA Glucuronic acid - Kav Elution volume relative to those of Blue Dextran (Kav=0) and glucose (Kav=1) - TFA Trifluoroacetic acid - V0 Void volume (centre of elution peak of Blue Dextran) - Vi Totally included volume (centre of elution peak of glucose) - XEH Xyloglucan endohydrolase (activity) - XET Xyloglucan endotransglucosylase (activity) - XLLGol A xyloglucan-derived oligosaccharide, xylose3·glucose3·galactose2·glucitol - XTH Xyloglucan endotransglucosylase/hydrolase (protein) - µ Ionic strength |
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Keywords: | Arabinogalactan-protein Cell wall restructuring Ionic strength Transglycosylation Wall polysaccharide Xyloglucan |
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