Skip to main content
PMC home page
Plant Physiology logoLink to Plant Physiology
. 1993 Jun;102(2):587–593. doi: 10.1104/pp.102.2.587

Respiration of Sugars in Spinach (Spinacia oleracea), Maize (Zea mays), and Chlamydomonas reinhardtii F-60 Chloroplasts with Emphasis on the Hexose Kinases.

K K Singh 1, C Chen 1, D K Epstein 1, M Gibbs 1
PMCID: PMC158816  PMID: 12231848

Abstract

The role of hexokinase in carbohydrate degradation in isolated, intact chloroplasts was evaluated. This was accomplished by monitoring the evolution of 14CO2 from darkened spinach (Spinacia oleracea), maize (Zea mays) mesophyll, and Chlamydomonas reinhardtii chloroplasts externally supplied with 14C-labeled fructose, glucose, mannose, galactose, maltose, and ribose. Glucose and ribose were the preferred substrates with the Chlamydomonas and maize chloroplasts, respectively. The rate of CO2 release from fructose was about twice that from glucose in the spinach chloroplast. Externally supplied ATP stimulated the rate of CO2 release. The pH optimum for CO2 release was 7.5 with ribose and fructose and 8.5 with glucose as substrates. Probing the outer membrane polypeptides of the intact spinach chloroplast with two proteases, trypsin and thermolysin, decreased 14CO2 release from glucose about 50% but had little effect when fructose was the substrate. Tryptic digestion decreased CO2 release from glucose in the Chlamydomonas chloroplast about 70%. 14CO2 evolution from [1-14C]-glucose-6-phosphate in both chloroplasts was unaffected by treatment with trypsin. Enzymic analysis of the supernatant (stroma) of the lysed spinach chloroplast indicated a hexokinase active primarily with fructose but with some affinity for glucose. The pellet (membranal fraction) contained a hexokinase utilizing both glucose and fructose but with considerably less total activity than the stromal enzyme. Treatment with trypsin and thermolysin eliminated more than 50% of the glucokinase activity but had little effect on fructokinase activity in the spinach chloroplast. Tryptic digestion of the Chlamydomonas chloroplast resulted in a loss of about 90% of glucokinase activity.

Full Text

The Full Text of this article is available as a PDF (739.7 KB).

Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. Chen C., Gibbs M. Coupling of Carbon Dioxide Fixation to the Oxyhydrogen Reaction in the Isolated Chloroplast of Chlamydomonas reinhardtii. Plant Physiol. 1992 Nov;100(3):1361–1365. doi: 10.1104/pp.100.3.1361. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Cline K., Andrews J., Mersey B., Newcomb E. H., Keegstra K. Separation and characterization of inner and outer envelope membranes of pea chloroplasts. Proc Natl Acad Sci U S A. 1981 Jun;78(6):3595–3599. doi: 10.1073/pnas.78.6.3595. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Cline K., Werner-Washburne M., Andrews J., Keegstra K. Thermolysin is a suitable protease for probing the surface of intact pea chloroplasts. Plant Physiol. 1984 Jul;75(3):675–678. doi: 10.1104/pp.75.3.675. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Heldt H. W. Adenine nucleotide translocation in spinach chloroplasts. FEBS Lett. 1969 Sep;5(1):11–14. doi: 10.1016/0014-5793(69)80280-2. [DOI] [PubMed] [Google Scholar]
  5. Heldt H. W., Chon C. J., Maronde D. Role of orthophosphate and other factors in the regulation of starch formation in leaves and isolated chloroplasts. Plant Physiol. 1977 Jun;59(6):1146–1155. doi: 10.1104/pp.59.6.1146. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Heldt H. W., Rapley L. Specific transport of inorganic phosphate, 3-phosphoglycerate and dihydroxyacetonephosphate, and of dicarboxylates across the inner membrane of spinach chloroplasts. FEBS Lett. 1970 Oct 5;10(3):143–148. doi: 10.1016/0014-5793(70)80438-0. [DOI] [PubMed] [Google Scholar]
  7. Klein U., Chen C., Gibbs M. Photosynthetic Properties of Chloroplasts from Chlamydomonas reinhardii. Plant Physiol. 1983 Jun;72(2):488–491. doi: 10.1104/pp.72.2.488. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Levi C., Gibbs M. Starch Degradation in Synchronously Grown Chlamydomonas reinhardtii and Characterization of the Amylase. Plant Physiol. 1984 Mar;74(3):459–463. doi: 10.1104/pp.74.3.459. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Levi C., Preiss J. Amylopectin degradation in pea chloroplast extracts. Plant Physiol. 1978 Feb;61(2):218–220. doi: 10.1104/pp.61.2.218. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. MACLACHLAN G. A., PORTER H. K. Replacement of oxidation by light as the energy source for glucose metabolism in tobacco leaf. Proc R Soc Lond B Biol Sci. 1959 Sep 1;150:460–473. doi: 10.1098/rspb.1959.0035. [DOI] [PubMed] [Google Scholar]
  11. Miernyk J. A., Dennis D. T. Mitochondrial, plastid, and cytosolic isozymes of hexokinase from developing endosperm of Ricinus communis. Arch Biochem Biophys. 1983 Oct 15;226(2):458–468. doi: 10.1016/0003-9861(83)90315-6. [DOI] [PubMed] [Google Scholar]
  12. Okita T. W., Greenberg E., Kuhn D. N., Preiss J. Subcellular localization of the starch degradative and biosynthetic enzymes of spinach leaves. Plant Physiol. 1979 Aug;64(2):187–192. doi: 10.1104/pp.64.2.187. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Schäfer G., Heber U. Glucose transport into spinach chloroplasts. Plant Physiol. 1977 Aug;60(2):286–289. doi: 10.1104/pp.60.2.286. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Singh K. K., Chen C., Gibbs M. Characterization of an Electron Transport Pathway Associated with Glucose and Fructose Respiration in the Intact Chloroplasts of Chlamydomonas reinhardtii and Spinach. Plant Physiol. 1992 Sep;100(1):327–333. doi: 10.1104/pp.100.1.327. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Stitt M., Bulpin P. V., ap Rees T. Pathway of starch breakdown in photosynthetic tissues of Pisum sativum. Biochim Biophys Acta. 1978 Nov 15;544(1):200–214. doi: 10.1016/0304-4165(78)90223-4. [DOI] [PubMed] [Google Scholar]
  16. Willeford K. O., Gibbs M. Localization of the Enzymes Involved in the Photoevolution of H(2) from Acetate in Chlamydomonas reinhardtii. Plant Physiol. 1989 Jul;90(3):788–791. doi: 10.1104/pp.90.3.788. [DOI] [PMC free article] [PubMed] [Google Scholar]

Articles from Plant Physiology are provided here courtesy of Oxford University Press

RESOURCES