Purification, characterization, and submitochondrial localization of a 58-kilodalton NAD(P)H dehydrogenase

Michael H. Luethy, Jay J. Thelen, Andrew F. Knudten, Thomas E. Elthon

Research output: Contribution to journalArticle

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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(β-aminoethyl ether)-N,N′-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%).

Original languageEnglish (US)
Pages (from-to)443-450
Number of pages8
JournalPlant physiology
Volume107
Issue number2
DOIs
StatePublished - Feb 1995

Fingerprint

NAD(P)H dehydrogenase (quinone)
NAD
Oxidoreductases
Proteins
proteins
Beta vulgaris
Zea mays
Mitochondria
mitochondria
beets
polyclonal antibodies
Antibodies
Mersalyl
p-Chloromercuribenzoic Acid
Platanus occidentalis
Flavin Mononucleotide
NADH Dehydrogenase
Mitochondrial Proteins
flavins
Mitochondrial Membranes

ASJC Scopus subject areas

  • Physiology
  • Genetics
  • Plant Science

Cite this

Purification, characterization, and submitochondrial localization of a 58-kilodalton NAD(P)H dehydrogenase. / Luethy, Michael H.; Thelen, Jay J.; Knudten, Andrew F.; Elthon, Thomas E.

In: Plant physiology, Vol. 107, No. 2, 02.1995, p. 443-450.

Research output: Contribution to journalArticle

Luethy, Michael H. ; Thelen, Jay J. ; Knudten, Andrew F. ; Elthon, Thomas E. / Purification, characterization, and submitochondrial localization of a 58-kilodalton NAD(P)H dehydrogenase. In: Plant physiology. 1995 ; Vol. 107, No. 2. pp. 443-450.
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