Abstract
HVPE is an excellent and often overlooked method for obtaining objective and meaningful information about cell-wall “building blocks” and their metabolic precursors. It provides not only a means of analysis of known compounds but also an insight into the charge and/or mass of any unfamiliar compounds that may be encountered. It can be used preparatively or analytically. It can achieve either “class separations” (e.g. delivering all hexose monophosphates into a single pool) or the resolution of different compounds within a given class (e.g. ADP-Glc from UDP-Glc; or GlcA from GalA). All information from HVPE about charge and mass can be obtained on minute traces of analytes, especially those that have been radiolabelled, e.g. by in-vivo feeding of a 3H- or 14C-labelled precursor. HVPE does not usually damage the substance under investigation (unless staining is used), so samples of interest can be eluted intact from the paper ready for further analysis. Although HVPE is a technique that has been available for several decades, recently it has tended to be sidelined, possibly because the apparatus is not widely available. Interested scientists are invited to contact the author about the possibility of accessing the Edinburgh apparatus.
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Notes
- 1.
i.e. a compound with 6 carbon atoms and two negative charges.
References
Offord, R. E. (1966). Electrophoretic mobilities of peptides on paper and their use in the determination of amide groups. Nature 211, 591–593.
Fry, S. C. (2000). The Growing Plant Cell Wall: Chemical and Metabolic Analysis, Reprint Edition. The Blackburn Press, Caldwell, NJ, pp. xviii + 333 [ISBN 1-930665-08-3].
Green, M. A., and Fry, S. C. (2005) Vitamin C degradation in plant cells via enzymatic hydrolysis of 4-O-oxalyl-l-threonate. Nature 433, 83–88.
Eshdat, Y., and Mirelman, D. (1972). An improved method for the recovery of compounds from paper chromatograms. Journal of Chromatography 65, 458–459.
Wright, K., and Northcote, D. H. (1975) An acidic oligosaccharide from maize slime. Phytochemistry 14, 1793–1798.
Popper, Z. A., Sadler, I. H., and Fry, S. C. (2003) α-d-Glucuronosyl-(1→3)-l-galactose, an unusual disaccharide from polysaccharides of the hornwort Anthoceros caucasicus. Phytochemistry 64, 325–335.
Sharples S. C., and Fry, S. C. (2007) Radio-isotope ratios discriminate between competing pathways of cell wall polysaccharide and RNA biosynthesis in living plant cells. Plant Journal 52, 252–262.
Kärkönen, A., and Fry, S. C. (2006) Novel characteristics of UDP-glucose dehydrogenase activities in maize: non-involvement of alcohol dehydrogenases in cell wall polysaccharide biosynthesis. Planta 223, 858–870.
Fry, S. C., Willis S. C., and Paterson, A. E. J. (2000). Intraprotoplasmic and wall-localised formation of arabinoxylan-bound diferulates and larger ferulate coupling-products in maize cell-suspension cultures. Planta 211, 679–692.
Piro, G., Perotto, S., Bonfante-Fasolo, P., and Dalessandro, G. (1988) Metabolism of d-(U-14C)glucosamine in seedlings of Calluna vulgaris (L) Hull. Journal of Plant Physiology 132, 695–701.
Lamport, D. T. A. (1967) Hydroxyproline-O-glycosidic linkage of plant cell plant cell wall glycoprotein extensin. Nature 216, 1322–1324.
Takahashi, T., Kakehi, J. -I. (2010) Polyamines: ubiquitous polycations with unique roles in growth and stress responses. Annals of Botany 105, 1–6
Lenucci M., Piro, G., Miller, J. G., Dalessandro, G., and Fry, S. C. (2005) Do polyamines contribute to plant cell wall assembly by forming amide bonds with pectins? Phytochemistry 66, 2581–2594.
Perrone P., Hewage, C., Sadler, I. H., and Fry, S. C. (1998). N α- and N ε-d-galacturonoyl-l-lysine amides: properties and possible occurrence in plant cell walls. Phytochemistry 49, 1879–90.
Kärkönen A., Warinowski, T., Teeri, T. H., Simola, L. K., and Fry, S. C. (2009) On the mechanism of apoplastic H2O2 production during lignin formation and elicitation in cultured spruce cells; peroxidases after elicitation. Planta 230, 553–567.
Weigel, H. (1963). Paper electrophoresis of carbohydrates. Advances in Carbohydrate Chemistry 18, 61–96.
Dumville J.C., and Fry, S. C. (2003) Gentiobiose: a novel oligosaccharin in ripening tomato fruit. Planta 216, 484–495.
Narasimham, S., Harpaz, N., Longmore, G., Carver, J. P., Grey, A. A., and Schachter, H. (1980) Control of glycoprotein synthesis: the purification by preparative paper electrophoresis in borate of glycopeptides containing high mannose and complex oligosaccharide chains linked to asparagine. Journal of Biological Chemistry 255, 4876–4884.
O’Looney, N., and Fry, S. C. (2005) Oxaziclomefone, a new herbicide, inhibits wall expansion in maize cell-cultures without affecting polysaccharide biosynthesis, xyloglucan transglycosylation, peroxidase action or apoplastic ascorbate oxidation. Annals of Botany 96, 1097–1107.
Wende, G., and Fry, S. C. (1996). 2-O-β-d-Xylopyranosyl-(5-O-feruloyl)-l-arabinose, a widespread component of grass cell walls. Phytochemistry 44, 1019–1030.
Miller J. G., Farkaš, V., Sharples, S. C., and Fry, S. C. (2007) O-Oligosaccharidyl-1-amino-1-deoxyalditols as intermediates for fluorescent labelling of oligosaccharides. Carbohydrate Research 342, 44–54.
Acknowledgements
I thank numerous past and present colleagues and students for electrophoretic data and experience presented in this chapter, and the UK BBSRC for financial support.
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Fry, S.C. (2011). High-Voltage Paper Electrophoresis (HVPE) of Cell-Wall Building Blocks and Their Metabolic Precursors. In: Popper, Z. (eds) The Plant Cell Wall. Methods in Molecular Biology, vol 715. Humana Press, Totowa, NJ. https://doi.org/10.1007/978-1-61779-008-9_4
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