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Glucan-phosphorylase forms in cotyledons of Pisum sativum L.: Localization, developmental change, in-vitro translation, and processing

Planta volume 185, pages 432–439 (1991)Cite this article

Abstract

The occurrence, location, and biosynthesis of glucan-phosphorylase (EC 2.4.1.1) isoenzymes were studied in cotyledons of developing or germinating seeds of Pisum sativum L. Type-I and type-II isoenzymes were detected, and were also localized by indirect immunofluorescence using polyclonal anti-type-I or anti-type-II phosphorylase antibodies. Type-I isoenzyme was found in the cytosol of parenchyma cells whereas the type-II enzyme form is a plastid protein which resides either in amyloplasts (in developing seeds) or in proplastids (in germinating seeds). During seed development, type-II phosphorylase was the predominant isoenzyme and the type-I isoenzyme represented a very minor compound. During germination, the latter increased whilst type-II phosphorylase remained at a constant level. In in-vitro translation experiments, type-I isoenzyme was observed as a final-size product with an apparent molecular weight of approx. 90 kDa. In contrast, type-II phosphorylase was translated as a high-molecular-weight precursor (116 kDa) which, when incubated with a stromal fraction of isolated intact pea chloroplasts, was processed to the size of the mature protein (105 kDa).

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Abbreviations

IgG:

immunoglobulin G

kDa:

kilodalton

poly(A)+ RNA:

polyadenylated RNA

SDS-PAGE:

sodium dodecyl sulfate-polyacrylamide gel electrophoresis

References

  • Anderson, D.J., Blobel, G. (1983) Immunoprecipitation of proteins from cell-free translations. Methods Enzymol. 96, 111–120

    Google Scholar 

  • Brisson, N., Giroux, H., Zollinger, M., Camirand, A., Simard, C. (1989) Maturation and subcellular compartmentation of potato starch phosphorylase. Plant Cell 1, 559–566

    Google Scholar 

  • Cashmore, A.R. (1982) The isolation of polyA+ messenger RNA from higher plants. In: Methods in chloroplast molecular biology, pp. 387–392, Edelman, M., Hallick, R.B., Chua, N.-H., eds. Elsevier Biomedical Press, Amsterdam

    Google Scholar 

  • Chua, N.-H., Schmidt, G.W. (1978) Post-translational transport into intact chloroplasts of a precursor to the small subunit of ribulose-1,5-bisphosphate carboxylase. Proc. Natl. Acad. Sci. USA 75, 6110–6114

    Google Scholar 

  • Conary, J., Nauerth, A., Burns, G., Hasilik, A., von Figura, K. (1986) Steroid sulfatase. Biosynthesis and processing in normal and mutant fibroblasts. Eur. J. Biochem. 158, 71–76

    Google Scholar 

  • Conrads, J., van Berkel, J., Schächtele, C., Steup, M., (1986) Non-hloroplast α-1,4-glucan phosphorylase from pea leaves: characterization and in situ localization by indirect immuno-fluorescence. Biochim. Biophys. Acta 882, 452–463

    Google Scholar 

  • Gupta, S.C., Beevers, L. (1987) Regulation of nitrite reductase. Cell-free translation and processing. Plant Physiol. 83, 750–754

    Google Scholar 

  • Hansen, S.A. (1988) Immunostaining of rocket immunoelectrophoresis precipitates too weak for identification following staining with Coomassie Brilliant Blue. Electrophoresis 9, 101–102

    Google Scholar 

  • Hershey, H.P., Quail, P.H. (1986) Identification of cDNA clones representing phytochrome and other low abundance red-light regulated sequences. Methods Enzymol. 118, 369–383

    Google Scholar 

  • Laemmli, U.K. (1970) Cleavage of structural proteins during the assembly of the head of bacteriophage T 4. Nature 227, 680–685

    PubMed  Google Scholar 

  • Nakano, K., Fukui T. (1986) The complete amino acid sequence of potato α-glucan phosphorylase. J. Biol. Chem. 261, 8230–8236

    Google Scholar 

  • Newgard, C.B., Hwang, P.K., Fletterick, R.J. (1989) The family of glycogen phosphorylases: structure and function. Crit. Rev. Biochem. Mol. Biol. 24, 69–98

    Google Scholar 

  • Palm, D., Goerl, P., Burger, K.J. (1985) Evolution of catalytic and regulatory sites in phosphorylases. Nature 313, 500–502

    Google Scholar 

  • Rittenhouse, J., Marcus, F. (1984) Peptide mapping by polyacrylamide gel electrophoresis after cleavage at aspartyl-prolyl peptide bonds in sodium dodecyl sulfate-containing buffers. Anal. Biochem. 138, 442–448

    Google Scholar 

  • Robinson, C., Ellis, R.J. (1984) Transport of proteins into chloroplasts. Partial purification of a chloroplast protease involved in the processing of imported precursor polypeptides. Eur. J. Biochem. 142, 337–342

    Google Scholar 

  • Schächtele, C., Steup, M. (1986) α-1,4-Glucan phosphorylase forms from leaves of spinach (Spinacia oleracea L.). I. In situ localization by indirect immunofluorescence. Planta 167, 444–451

    Google Scholar 

  • Schneider, E.M., Becker, J.-U., Volkmann, D. (1981) Biochemical properties of potato phosphorylase change with its intracellular localization as revealed by immunological methods. Planta 151, 124–134

    Google Scholar 

  • Shimomura, S., Nagai, M., Fukui, T. (1982) Comparative glucan specificities of two types of spinach leaf phosphorylase. J. Biochem. 91, 703–717

    Google Scholar 

  • Smith, A.M., Quinton-Tulloch, J., Denyer, K. (1990) Characteristics of plastids responsible for starch synthesis in developing pea embryos. Planta 180, 517–523

    Google Scholar 

  • Steup, M. (1988) Starch degradation. Biochemistry of plants, vol. 14: Carbohydrates, pp. 255–296, Preiss, J., ed. Academic Press, New York

    Google Scholar 

  • Steup, M. (1990) Starch degrading enzymes. In: Methods in plant biochemistry, pp. 103–128, Lea, P.J., ed. Academic Press, New York

    Google Scholar 

  • Steup, M., Latzko, E. (1979) Intracellular localization of phosphorylases in spinach and pea leaves. Planta 145, 69–75

    Google Scholar 

  • Steup, M., Schächtele, C. (1981) Mode of glucan degradation by purified phosphorylase forms from spinach leaves. Planta 153, 351–361

    Google Scholar 

  • Steup, M., Schächtele, C., Melkonian, M. (1986) Light-mediated changes in the plastidic phosphorylase patterns in shoots of Pisum sativum. Physiol. Plant. 66, 234–244

    Google Scholar 

  • Towbin, H., Staehelin, T., Gordon, J. (1979) Electrophoretic transfer of proteins from polyacrylamide gels to nitrocellulose sheets: procedure and some applications. Proc. Natl. Acad. Sci. USA 76, 4350–4354

    Google Scholar 

  • Viola, R., Davies, H.V., Chudek, A.R. (1991) Pathways of starch and sucrose biosynthesis in developing tubers of potato (Sola- num tuberosum L.) and seeds of faba bean (Vicia faba L.). Elucidation by 13C-nuclear-magnetic-resonance spectroscopy. Planta 183, 202–208

    Google Scholar 

  • von Figura, K., Gieselmann, V., Hasilik, A. (1985) Mannose-6-phosphate-specific receptor is a transmembrane protein with a C-terminal extension oriented towards the cytosol. Biochem. J. 225, 543–547

    Google Scholar 

  • Westhoff, P., Zetsche, K. (1981) Regulation of the synthesis of ribulose-1,5-bisphosphate carboxylase and its subunits in the flagellate Chlorogonium elongatum. Eur. J. Biochem. 116, 261–267

    Google Scholar 

  • Yang, Y., Steup, M. (1990) Polysaccharide fraction from higher plants which strongly interacts with the cytosolic phosphorylase isozyme. I. Isolation and characterization. Plant Physiol. 94, 960–969

    Google Scholar 

  • Yu, F., Jen, Takeuchi, E., Inouye, M., Nakayama, H., Tagaya, M., Fukui, T. (1988) α-Glucan phosphorylase from Escherichia coli, Cloning of the gene, and purification and characterization of the protein. J. Biol. Chem. 263, 13706–13711

    Google Scholar 

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Author notes

  1. Joachim van Berkel

    Present address: Max-Planck-Institut für Züchtungsforschung, Carl-von-Linné-Weg 10, 30, Köln, Germany

  2. Jutta Conrads-Strauch

    Present address: John Innes Institute, Colney Lane, NR4 7UH, Norwich, UK

Affiliations

  1. Institut für Botanik, Westfälische Wilhelms-Universität Münster, Schlossgarten 3, W-4400, Münster, Germany

    Joachim van Berkel, Jutta Conrads-Strauch & Martin Steup

Authors
  1. Joachim van Berkel
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  2. Jutta Conrads-Strauch
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  3. Martin Steup
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Additional information

This work has been made possible by grants from the Deutsche Forschungsgemeinschaft. The authors are endebted to Mrs. Karin Niehüser for help in the immunocytochemical studies.

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van Berkel, J., Conrads-Strauch, J. & Steup, M. Glucan-phosphorylase forms in cotyledons of Pisum sativum L.: Localization, developmental change, in-vitro translation, and processing. Planta 185, 432–439 (1991). https://doi.org/10.1007/BF00201068

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  • DOI: https://doi.org/10.1007/BF00201068

Key words

  • Glucan phosphorylase
  • Isoenzyme processing
  • Pisum
  • Starch metabolism