Planta

, Volume 199, Issue 2, pp 209–218

Starch debranching enzyme (R-enzyme or pullulanase) from developing rice endosperm: purification, cDNA and chromosomal localization of the gene

  • Yasunori Nakamura
  • Takayuki Umemoto
  • Naoki Ogata
  • Yoshihide Kuboki
  • Masahiro Yano
  • Takuji Sasaki
Article

DOI: 10.1007/BF00196561

Cite this article as:
Nakamura, Y., Umemoto, T., Ogata, N. et al. Planta (1996) 199: 209. doi:10.1007/BF00196561

Abstract

Starch debranching enzyme (R-enzyme or pullulanase) was purified to homogeneity from developing endosperm of rice (Oryza sativa L. cv. Fujihikari) using a variety of high-performance liquid chromatography columns, and characterized. A cDNA clone encoding the full length of the rice endosperm debranching enzyme was isolated and its nucleotide sequence was determined. The cDNA contains an open reading frame of 2958 bp. The mature debranching enzyme of rice appears to be composed of 912 amino acids with a predicted relative molecular mass (Mr) of 102069 Da, similar in size to its Mr of about 100 000 Da estimated by polyacrylamide gel electrophoresis in sodium dodecyl sulfate. The amino acid sequence of rice debranching enzyme is substantially similar to that of bacterial pullulanase, while it bears little similarity to that of bacterial isoamylase or to glycogen debranching enzymes from human muscle and rabbit muscle. Southern blot analyses strongly suggest that the debranching enzyme gene is present as a single copy in the rice genome. Analysis by restriction fragment length polymorphism with a probe including the 3′-untranslated region of cDNA for rice debranching enzyme confirmed that the debranching enzyme gene is located on chromosome 4.

Key words

AmylopectinDebranching enzymeGlycogenOryza endospermPullulanaseR-enzymeStarch

Abbreviations

RFLP

restriction fragment length polymorphism

SDS-PAGE

polyacrylamide gel electrophoresis in sodium dodecyl sulfate

Copyright information

© Springer-Verlag 1996

Authors and Affiliations

  • Yasunori Nakamura
    • 1
  • Takayuki Umemoto
    • 2
  • Naoki Ogata
    • 1
  • Yoshihide Kuboki
    • 3
  • Masahiro Yano
    • 3
  • Takuji Sasaki
    • 3
  1. 1.Department of Applied PhysiologyNational Institute of Agrobiological Resources (NIAR)IbarakiJapan
  2. 2.Department of Lowland FarmingTohoku National Agricultural Experiment StationAkitaJapan
  3. 3.Rice Genome Research Program, NIAR/STAFFIbarakiJapan