Skip to main content
PMC home page
Plant Physiology logoLink to Plant Physiology
. 1995 Feb;107(2):443–450. doi: 10.1104/pp.107.2.443

Purification, Characterization, and Submitochondrial Localization of a 58-Kilodalton NAD(P)H Dehydrogenase.

M H Luethy 1, J J Thelen 1, A F Knudten 1, T E Elthon 1
PMCID: PMC157146  PMID: 12228370

Abstract

An NADH dehydrogenase activity from red beet (Beta vulgaris L.) root mitochondria was purified to a 58-kD protein doublet. An immunologically related dehydrogenase was partially purified from maize (Zea mays L. B73) mitochondria to a 58-kD protein doublet, a 45-kD protein, and a few other less prevalent proteins. Polyclonal antibodies prepared against the 58-kD protein of red beet roots were found to immunoprecipitate the NAD(P)H dehydrogenase activity. The antibodies cross-reacted to similar proteins in mitochondria from a number of plant species but not to rat liver mitochondrial proteins. The polyclonal antibodies were used in conjunction with maize mitochondrial fractionation to show that the 58-kD protein was likely part of a protein complex loosely associated with the membrane fraction. A membrane-impermeable protein cross-linking agent was used to further show that the majority of the 58-kD protein was located on the outer surface of the inner mitochondrial membrane or in the intermembrane space. Analysis of the cross-linked 58-kD NAD(P)H dehydrogenase indicated that specific proteins of 64, 48, and 45 kD were cross-linked to the 58-kD protein doublet. The NAD(P)H dehydrogenase activity was not affected by ethyleneglycol-bis([beta]-aminoethyl ether)-N,N[prime] -tetraacetic acid or CaCl2, was stimulated somewhat (21%) by flavin mononucleotide, was inhibited by p-chloromercuribenzoic acid (49%) and mersalyl (40%), and was inhibited by a bud scale extract of Platanus occidentalis L. containing platanetin (61%).

Full Text

The Full Text of this article is available as a PDF (2.3 MB).

Selected References

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

  1. Chauveau M., Lance C. Purification and Partial Characterization of Two Soluble NAD(P)H Dehydrogenases from Arum maculatum Mitochondria. Plant Physiol. 1991 Mar;95(3):934–942. doi: 10.1104/pp.95.3.934. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Coleman J. O.D., Palmer J. M. Role of Ca(2+) in the oxidation of exogenous NADH by plant mitochondria. FEBS Lett. 1971 Oct 1;17(2):203–208. doi: 10.1016/0014-5793(71)80148-5. [DOI] [PubMed] [Google Scholar]
  3. Cook N. D., Cammack R. Purification and characterization of the rotenone-insensitive NADH dehydrogenase of mitochondria from Arum maculatum. Eur J Biochem. 1984 Jun 15;141(3):573–577. doi: 10.1111/j.1432-1033.1984.tb08231.x. [DOI] [PubMed] [Google Scholar]
  4. Cottingham I. R., Cleeter M. W., Ragan C. I., Moore A. L. Immunological analysis of plant mitochondrial NADH dehydrogenases. Biochem J. 1986 May 15;236(1):201–207. doi: 10.1042/bj2360201. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Cottingham I. R., Moore A. L. Partial purification and properties of the external NADH dehydrogenase from cuckoo-pint (Arum maculatum) mitochondria. Biochem J. 1984 Nov 15;224(1):171–179. doi: 10.1042/bj2240171. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Douce R., Mannella C. A., Bonner W. D., Jr The external NADH dehydrogenases of intact plant mitochondria. Biochim Biophys Acta. 1973 Jan 18;292(1):105–116. doi: 10.1016/0005-2728(73)90255-7. [DOI] [PubMed] [Google Scholar]
  7. Elthon T. E., McIntosh L. Characterization and Solubilization of the Alternative Oxidase of Sauromatum guttatum Mitochondria. Plant Physiol. 1986 Sep;82(1):1–6. doi: 10.1104/pp.82.1.1. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Elthon T. E., Nickels R. L., McIntosh L. Monoclonal antibodies to the alternative oxidase of higher plant mitochondria. Plant Physiol. 1989 Apr;89(4):1311–1317. doi: 10.1104/pp.89.4.1311. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Hayes M. K., Luethy M. H., Elthon T. E. Mitochondrial malate dehydrogenase from corn : purification of multiple forms. Plant Physiol. 1991 Dec;97(4):1381–1387. doi: 10.1104/pp.97.4.1381. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Klein R. R., Burke J. J. Separation Procedure and Partial Characterization of Two NAD(P)H Dehydrogenases from Cauliflower Mitochondria. Plant Physiol. 1984 Oct;76(2):436–441. doi: 10.1104/pp.76.2.436. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Krömer S., Heldt H. W. On the Role of Mitochondrial Oxidative Phosphorylation in Photosynthesis Metabolism as Studied by the Effect of Oligomycin on Photosynthesis in Protoplasts and Leaves of Barley (Hordeum vulgare). Plant Physiol. 1991 Apr;95(4):1270–1276. doi: 10.1104/pp.95.4.1270. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Larson E., Howlett B., Jagendorf A. Artificial reductant enhancement of the Lowry method for protein determination. Anal Biochem. 1986 Jun;155(2):243–248. doi: 10.1016/0003-2697(86)90432-x. [DOI] [PubMed] [Google Scholar]
  13. Luethy M. H., Hayes M. K., Elthon T. E. Partial Purification and Characterization of Three NAD(P)H Dehydrogenases from Beta vulgaris Mitochondria. Plant Physiol. 1991 Dec;97(4):1317–1322. doi: 10.1104/pp.97.4.1317. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Ravanel P., Tissut M., Douce R. Platanetin: A Potent Natural Uncoupler and Inhibitor of the Exogenous NADH Dehydrogenase in Intact Plant Mitochondria. Plant Physiol. 1986 Feb;80(2):500–504. doi: 10.1104/pp.80.2.500. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Wilson R. H., Hanson J. B. The effect of respiratory inhibitors on NADH, succinate and malate oxidation in corn mitochondria. Plant Physiol. 1969 Sep;44(9):1335–1341. doi: 10.1104/pp.44.9.1335. [DOI] [PMC free article] [PubMed] [Google Scholar]

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

RESOURCES