• We are sorry, but NCBI web applications do not support your browser and may not function properly. More information
Logo of pnasPNASInfo for AuthorsSubscriptionsAboutThis Article
Proc Natl Acad Sci U S A. Oct 15, 1993; 90(20): 9441–9445.
PMCID: PMC47584

Ethylene induces de novo synthesis of chlorophyllase, a chlorophyll degrading enzyme, in Citrus fruit peel.

Abstract

Chlorophyllase (Chlase; EC 3.1.1.14) was extracted from plastid fractions of ethylene-treated orange fruit peel and purified 400-fold to homogeneity by gel filtration, hydrophobic chromatography, and preparative SDS/PAGE of nonheated protein. SDS/PAGE of nonheated purified enzyme indicated that Chlase activity is associated with a single protein band migrating at an apparent molecular mass of 25 kDa whereas the heated purified enzyme had a molecular mass of 35 kDa. The N-terminal sequence of the purified protein was determined. The purified enzyme was used as an immunogen for raising antibodies in rabbits. The antiserum was highly specific and on Western blots recognized both the heated and the nonheated form of Chlase. The antibodies also recognized the solubilized enzyme, as shown by an immunoprecipitation assay and by antigen-antibody capture assays in microtiter plates. Treatment with ethylene, which enhances degreening, increased Chlase activity 12-fold. Immunoblot analyses of crude extracts from ethylene-treated fruit detected a strong signal of the Chlase protein, while only a trace level of the enzyme protein could be detected in air. Gibberellin A3 and N6-benzyladenine partly counteracted the ethylene-induced increase in Chlase activity as well as the immunodetected upsurge of the Chlase protein. Ethylene appears to enhance the degreening of citrus fruit through de novo synthesis of the Chlase protein, which in turn is inhibited by the senescence-delaying regulators, gibberellin A3 and N6-benzyladenine. The Chlase enzyme protein may, therefore, serve as a model system for studying the hormonal molecular regulation of fruit ripening and senescence.

Full text

Full text is available as a scanned copy of the original print version. Get a printable copy (PDF file) of the complete article (1.3M), or click on a page image below to browse page by page. Links to PubMed are also available for Selected References.

Images in this article

Selected References

These references are in PubMed. This may not be the complete list of references from this article.
  • Goldschmidt EE, Aharoni Y, Eilati SK, Riov JW, Monselise SP. Differential counteraction of ethylene effects by gibberellin a(3) and n(6)-benzyladenine in senescing citrus peel. Plant Physiol. 1977 Feb;59(2):193–195. [PMC free article] [PubMed]
  • Purvis AC, Barmore CR. Involvement of ethylene in chlorophyll degradation in peel of citrus fruits. Plant Physiol. 1981 Oct;68(4):854–856. [PMC free article] [PubMed]
  • Amir-Shapira D, Goldschmidt EE, Altman A. Chlorophyll catabolism in senescing plant tissues: In vivo breakdown intermediates suggest different degradative pathways for Citrus fruit and parsley leaves. Proc Natl Acad Sci U S A. 1987 Apr;84(7):1901–1905. [PMC free article] [PubMed]
  • Eilati SK, Goldschmidt EE, Monselise SP. Hormonal control of colour changes in orange peel. Experientia. 1969 Feb 15;25(2):209–210. [PubMed]
  • McFeeters RF, Chichester CO, Whitaker JR. Purification and Properties of Chlorophyllase from Ailanthus altissima (Tree-of-Heaven). Plant Physiol. 1971 May;47(5):609–618. [PMC free article] [PubMed]
  • Moran R, Porath D. Chlorophyll determination in intact tissues using n,n-dimethylformamide. Plant Physiol. 1980 Mar;65(3):478–479. [PMC free article] [PubMed]
  • Young PR. Enhancement of immunoblot staining using a mixed chromogenic substrate. J Immunol Methods. 1989 Jul 26;121(2):295–296. [PubMed]
  • Shioi Y, Sasa T. Purification of solubilized chlorophyllase from Chlorella protothecoides. Methods Enzymol. 1986;123:421–427. [PubMed]
  • Tanaka K, Kakuno T, Yamashita J, Horio T. Purification and properties of chlorophyllase from greened rye seedlings. J Biochem. 1982 Dec;92(6):1763–1773. [PubMed]
  • Reynolds JA, Tanford C. The gross conformation of protein-sodium dodecyl sulfate complexes. J Biol Chem. 1970 Oct 10;245(19):5161–5165. [PubMed]
  • Devereux J, Haeberli P, Smithies O. A comprehensive set of sequence analysis programs for the VAX. Nucleic Acids Res. 1984 Jan 11;12(1 Pt 1):387–395. [PMC free article] [PubMed]
  • Dellapenna D, Alexander DC, Bennett AB. Molecular cloning of tomato fruit polygalacturonase: Analysis of polygalacturonase mRNA levels during ripening. Proc Natl Acad Sci U S A. 1986 Sep;83(17):6420–6424. [PMC free article] [PubMed]
  • Dostal HC, Leopold AC. Gibberellin delays ripening of tomatoes. Science. 1967 Dec 22;158(3808):1579–1580. [PubMed]
  • Tucker ML, Sexton R, Del Campillo E, Lewis LN. Bean abscission cellulase : characterization of a cDNA clone and regulation of gene expression by ethylene and auxin. Plant Physiol. 1988 Dec;88(4):1257–1262. [PMC free article] [PubMed]
  • Gray J, Picton S, Shabbeer J, Schuch W, Grierson D. Molecular biology of fruit ripening and its manipulation with antisense genes. Plant Mol Biol. 1992 May;19(1):69–87. [PubMed]
  • Theologis A, Zarembinski TI, Oeller PW, Liang X, Abel S. Modification of fruit ripening by suppressing gene expression. Plant Physiol. 1992 Oct;100(2):549–551. [PMC free article] [PubMed]

Articles from Proceedings of the National Academy of Sciences of the United States of America are provided here courtesy of National Academy of Sciences

Formats:

Related citations in PubMed

See reviews...See all...

Cited by other articles in PMC

See all...

Links

  • Gene
    Gene
    Gene links
  • MedGen
    MedGen
    Related information in MedGen
  • Nucleotide
    Nucleotide
    Published Nucleotide sequences
  • Protein
    Protein
    Published protein sequences
  • PubMed
    PubMed
    PubMed citations for these articles

Recent Activity

Your browsing activity is empty.

Activity recording is turned off.

Turn recording back on

See more...