Jump to ContentJump to Main Navigation
Show Summary Details
More options …


12 Issues per year

See all formats and pricing
More options …
Volume 64, Issue 2


Oligogalacturonate hydrolase with unique substrate preference from the pulp of parsley roots

Dana Flodrová
  • Faculty of Chemistry, Technical University of Brno, Purkyňova 118, CZ-61200, Brno, Czech Republic
  • Institute of Analytical Chemistry, v.v.i., Academy of Sciences of the Czech Republic, Veveří 97, CZ-60200, Brno, Czech Republic
  • Email
  • Other articles by this author:
  • De Gruyter OnlineGoogle Scholar
/ Soňa Garajová / Anna Malovíková / Danica Mislovičová / Jiřina Omelková / Eva Stratilová
Published Online: 2009-02-20 | DOI: https://doi.org/10.2478/s11756-009-0038-2


The main form of pectate hydrolases in the cell wall of parsley roots showed a unique substrate preference of a plant exopolygalacturonase because it clearly preferred the substrates with degree of polymerization about 10. This form was separated from the others, purified and characterized. Enzyme exhibited sharp pH optimum corresponding to pH 4.7, molecular mass 53.5 kDa, and isoelectric point 5.3. It was stable at 50°C in 2-h assay and had optimum of temperature at 60°C (activation energy being 37.0 kJ/mol). The interaction with concanavalin A indicated the glycosylation of enzyme. Substrates were cleaved from the non-reducing end.

Keywords: exopolygalacturonase; oligogalacturonate hydrolase; Petroselim crispum

  • [1] Abbott D.W. & Boraston A.B. 2007. The structural basis for exopolygalacturonase activity in family 28 glycoside hydrolase. J. Mol. Biol. 368: 1215–1222. http://dx.doi.org/10.1016/j.jmb.2007.02.083CrossrefWeb of ScienceGoogle Scholar

  • [2] Biely P., Benen J., Heinrichová K., Kester H.C. & Visser J. 1996. Inversion of configuration during hydrolysis of α-1,4-galacturonic linkage by three Aspergillus polygalacturonases. FEBS Lett. 382: 249–255. http://dx.doi.org/10.1016/0014-5793(96)00171-8CrossrefGoogle Scholar

  • [3] Bradford M.M. 1976. A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein-dye binding. Anal. Biochem. 72: 248–254. http://dx.doi.org/10.1016/0003-2697(76)90527-3CrossrefGoogle Scholar

  • [4] Carpita N.C. & Gibeaut D.M. 1993. Structural models of primary cell walls in flowering plants: consistency of molecular structure with the physical properties of the walls during growth. Plant J. 3: 1–30. http://dx.doi.org/10.1111/j.1365-313X.1993.tb00007.xCrossrefGoogle Scholar

  • [5] Cho S.W., Lee S. & Shin W. 2001. The X-ray structure of Aspergillus aculeatus polygalacturonase and a modeled structure of the polygalacturonase-octagalacturonate complex. J. Mol. Biol. 314: 863–878. http://dx.doi.org/10.1006/jmbi.2001.4919CrossrefGoogle Scholar

  • [6] Dzúrová M., Linek K. & Stratilová E. 1995. Inhibition of exopolygalacturonase from carrots by reaction products and by their basic analogues. Biologia 50: 595–599. Google Scholar

  • [7] Federici L., Caprari C., Mattei B., Savino C., Di Matteo A., De Lorenzo G., Cervone F. & Tsernoglou D. 2001. Structural requirements of endopolygalacturonase for the interaction with PGIP. Proc. Natl. Acad. Sci. USA 98: 13425–13430. http://dx.doi.org/10.1073/pnas.231473698CrossrefGoogle Scholar

  • [8] Flodrová D., Dzúrová M., Lišková D., Ait Mohand F., Mislovičová D., Malovíková A., Voburka Z., Omelková J. & Stratilová, E. 2007. Pectate hydrolases of parsley (Petroselinum crispum) roots. Z. Naturforsch. 62c: 382–388. Google Scholar

  • [9] Garcia-Romera I. & Fry S.C. 1995. The longevity of biologicallyactive oligogalacturonides in rose cell cultures. Degradation by exo-polygalacturonase. J. Exp. Botany 46: 1853–1857. http://dx.doi.org/10.1093/jxb/46.12.1853CrossrefGoogle Scholar

  • [10] Hatanaka C. & Ozawa J. 1964. Enzymic degradation of pectic acid. I. Limited hydrolysis of pectic acids by carrot exopolygalacturonase. Agric. Biol. Chem. 28: 672–632. Google Scholar

  • [11] Hasegawa S. & Nagel C.W. 1968. Isolation of an oligogalacturonate hydrolase from a Bacillus specie. Arch. Biochem. Biophys. 124: 513–520. http://dx.doi.org/10.1016/0003-9861(68)90360-3CrossrefGoogle Scholar

  • [12] Heinrichová K. 1977. Isolation, characterization and mode of action of exo-d-galacturonanase from carrot. Collect. Czech. Chem. Commun. 42: 3214–3221. CrossrefGoogle Scholar

  • [13] Heinrichová K. 1983. Preparation of oligogalacturonic acids by enzymatic hydrolysis. Biologia 38: 335–342. Google Scholar

  • [14] Heinrichová K., Heinrich J., Dzúrová M. & Ziolecki A. 1993. Mode of action and partial purification of the active centre of exopoly-α-d-galacturonosidase from Selenomonas ruminantium. Collect. Czech. Chem. Commun. 58: 681–692. http://dx.doi.org/10.1135/cccc19930681CrossrefGoogle Scholar

  • [15] Heinrichová K. & Rexová-Benková L’. 1976. Purification and characterization of an extracellular exo-d-galacturonanase of Aspergillus niger. Biochim. Biophys. Acta 422: 349–356. Google Scholar

  • [16] Henrissat B. 1991. A classification of glycosyl hydrolases based on amino-acid-sequence similarities. Biochem. J. 280: 309–316. Google Scholar

  • [17] Henrissat B. & Davies G. 1997. Structural and sequence-based classification of glycoside hydrolases. Curr. Opin. Struct. Biol. 7: 637–644. http://dx.doi.org/10.1016/S0959-440X(97)80072-3CrossrefGoogle Scholar

  • [18] Kester H.C.M., Kusters-van Someren M.A., Müller Y. & Visser J. 1996. Primary structure and characterization of an exopolygalacturonase from Aspergillus tubigensis. Eur. J. Biochem. 240: 738–746. http://dx.doi.org/10.1111/j.1432-1033.1996.0738h.xCrossrefGoogle Scholar

  • [19] Kohn R. & Furda I. 1967. Calcium ion activity in solutions of calcium pectinate. Collect. Czech. Chem. Commun. 32: 1925–1937. CrossrefGoogle Scholar

  • [20] Kohn R. & Luknár O. 1977. Intermolecular calcium ion binding on polyuronates — polygalacturonate and polyguluronate. Collect. Czech. Chem. Commun. 42: 731–744. CrossrefGoogle Scholar

  • [21] Koller A. & Neukom H. 1964. Detection of oligogalacturonic acids by thin-layer chromatography. Biochim. Biophys. Acta 83: 366–367. Google Scholar

  • [22] Liao C.H., Revear L., Hotchkiss A. & Savary B. 1999. Genetic and biochemical characterization of an exopolygalacturonase and pectate lyase from Yersinia enterocolitica. Can. J. Microbiol. 45: 396–403. http://dx.doi.org/10.1139/cjm-45-5-396CrossrefGoogle Scholar

  • [23] Markovič O. & Janeček Š. 2001. Pectin degrading glycoside hydrolases of family 28: sequence-structural features, specificities and evolution. Protein Eng. 14: 615–631. http://dx.doi.org/10.1093/protein/14.9.615CrossrefGoogle Scholar

  • [24] Martens-Uzunova E.S., Zandleven J.S., Benen J.A.E., Awad H., Kools H.J., Beldman G., Voragen A.G.J., van den Berg J.A. & Schaap P.J. 2006. A new group of exo-acting family 28 glycoside hydrolases of Aspergillus niger that are involved in pectin degradation. Biochem. J. 400: 43–52. http://dx.doi.org/10.1042/BJ20060703CrossrefGoogle Scholar

  • [25] Musell H.W. & Strouse B. 1972. Characterization of two polygalacturonases produced by Verticillium albo-atrum. Can. J. Biochem. 50: 625–632. http://dx.doi.org/10.1139/o72-086CrossrefGoogle Scholar

  • [26] Niture S. 2008. Comparative biochemical and structural characterizations of fungal polygalacturonases. Biologia 63: 1–19. http://dx.doi.org/10.2478/s11756-008-0018-yWeb of ScienceCrossrefGoogle Scholar

  • [27] Parenicová L., Kester H.C.M., Benen J.A.E. & Visser J. 2000. Characterization of a novel endopolygalacturonase from Aspergillus niger with unique kinetic properties. FEBS Lett. 467: 333–336. http://dx.doi.org/10.1016/S0014-5793(00)01173-XCrossrefGoogle Scholar

  • [28] Pickersgill R., Smith D., Worboys K. & Jenkins J. 1998. Crystal structure of polygalacturonase from Erwinia carotovora ssp. carotovora. J. Biol. Chem. 273: 24660–24664. http://dx.doi.org/10.1074/jbc.273.38.24660CrossrefGoogle Scholar

  • [29] Pressey R. & Avants J.K. 1975. Modes of action of carrot and peach exopolygalacturonases. Phytochemistry 14: 957–961. http://dx.doi.org/10.1016/0031-9422(75)85166-1CrossrefGoogle Scholar

  • [30] Radola B.J. 1980. Ultrathin-layer isoelectric focusing in 50–100 μm polyacrylamide gels on silanized plates or polyester films. Electrophoresis 1: 43–56. http://dx.doi.org/10.1002/elps.1150010109CrossrefGoogle Scholar

  • [31] Rayle D.L. & Cleland R.E. 1992. The acid growth theory of auxin-induced cell elongation is alive and well. Plant Physiol. 99: 1271–1274. http://dx.doi.org/10.1104/pp.99.4.1271CrossrefGoogle Scholar

  • [32] Rexová-Benková L’. 1970. Separation of oligogalacturonic acids by dextran gel chromatography. Chem. Zvesti 24: 59–62. Google Scholar

  • [33] Rexová-Benková L’. & Markovič O. 1976. Pectic enzymes. Adv. Carbohydr. Chem. Biochem. 33: 323–385. http://dx.doi.org/10.1016/S0065-2318(08)60285-1CrossrefGoogle Scholar

  • [34] Shimizu T., Nakatsu T., Miyairi K., Okuno T. & Kato H. 2002. Active-site architecture of endopolygalacturonase I from Stereum purpurum revealed by crystal structures in native and ligand-bound forms at atomic resolution. Biochemistry 41: 6651–6659. http://dx.doi.org/10.1021/bi025541aCrossrefGoogle Scholar

  • [35] Somogyi M. 1952. Notes on sugar determination. J. Biol. Chem. 195: 19–23. Google Scholar

  • [36] Stratilová E., Dzúrová M., Breierová E. & Omelková J. 2006. Production and biochemical characterization of polygalacturonases produced by Aureobasidium pullulans from forest soil. Ann. Microbiol. 56: 35–40. http://dx.doi.org/10.1007/BF03174967CrossrefGoogle Scholar

  • [37] Stratilová E., Dzúrová M., Malovíková A. & Omelková J. 2005. Oligogalacturonate hydrolase from carrot roots. Z. Naturforsch. 60c: 899–905. Google Scholar

  • [38] Stratilová E., Mislovičová D. & Dzúrová M. 1996. Purification of exopolygalacturonase by affinity chromatography on concanavalin A — bead cellulose. Biotechnol. Tech. 10: 363–366. http://dx.doi.org/10.1007/BF00173256CrossrefGoogle Scholar

  • [39] Torki M., Mandaron P., Mache R. & Falconet D. 2000. Characterization of a ubiquitous expressed gene family encoding polygalacturonase in Arabidopsis thaliana. Gene 242: 427–436. http://dx.doi.org/10.1016/S0378-1119(99)00497-7CrossrefGoogle Scholar

  • [40] van Pouderoyen G., Snijder H.J., Bennen J.A. & Dijkstra B.W. 2003. Structural insights into the processivity of endopolygalacturonase I from Aspergillus niger. FEBS Lett. 554: 462–466. http://dx.doi.org/10.1016/S0014-5793(03)01221-3CrossrefGoogle Scholar

  • [41] Van Rijssel M., Smidt M.P., van Kouwen G. & Hansen T. 1993. Involvement of an intracellular oligogalacturonate hydrolase in metabolism of pectin by Clostridium thermosacchrolyticum. Appl. Environ. Microbiol. 59: 837–842. Google Scholar

  • [42] van Santen Y., Bennen J.A., Schroter K.H., Kalk K.H., Armand S., Visser J. & Dijkstra B.W. 1999. 1.68 Å crystal structure of endopolygalacturonase II from Aspergillus niger and identification of active site residues by site-directed mutagenesis. J. Biol. Chem. 274: 30474–30480. http://dx.doi.org/10.1074/jbc.274.43.30474CrossrefGoogle Scholar

  • [43] Vincken J.P., Schols H.A., Oomen R.J., Beldman G., Visser R.G.F. & Voragen A.G.J. 2003a. Pectin — the hairy thing, pp. 47–50. In: Voragen A.G.J., Schols H.A. & Visser R.G.F. (eds), Advances in Pectin and Pectinase Research, Kluwer Academic Publishers, Dordrecht. Google Scholar

  • [44] Vincken J.P., Schols H.A., Oomen R.J., McCann M.C., Ulvskov P., Voragen A.G.J. & Visser R.G.F. 2003b. If homogalacturonan were a side chain of rhamnogalacturonan I. Implications for cell wall architecture. Plant Physiol. 132: 1781–1789. http://dx.doi.org/10.1104/pp.103.022350CrossrefGoogle Scholar

  • [45] Voragen A.G.J., Pilnik W., Thibault J.F., Axelos M.A.V. & Renard C.M.G.C. 1995. Pectins, pp. 287–339. In: Stephen A.M. (ed.), Food Polysaccharides, Dekker, Inc., New York-Basel-Hong Kong. Google Scholar

  • [46] Wray W., Boulikas T., Wray V.P. & Hancock R. 1981. Silver staining of proteins in polyacrylamide gels. Anal. Biochem. 118: 197–203. http://dx.doi.org/10.1016/0003-2697(81)90179-2CrossrefGoogle Scholar

About the article

Published Online: 2009-02-20

Published in Print: 2009-04-01

Citation Information: Biologia, Volume 64, Issue 2, Pages 228–234, ISSN (Online) 1336-9563, ISSN (Print) 0006-3088, DOI: https://doi.org/10.2478/s11756-009-0038-2.

Export Citation

© 2009 Slovak Academy of Sciences. This work is licensed under the Creative Commons Attribution-NonCommercial-NoDerivatives 3.0 License. BY-NC-ND 3.0

Comments (0)

Please log in or register to comment.
Log in