Partial purification and characterization of three NAD(P)H dehydrogenases from Beta vulgaris mitochondria

Michael H. Luethy, Marianne K. Hayes, Thomas E Elthon

Research output: Contribution to journalArticle

26 Citations (Scopus)

Abstract

Mitochondria isolated from the taproot of beet (Beta vulgaris) were used in an effort to identify and partially purify the proteins constituting the exogenous NADH dehydrogenase. Three NAD(P)H dehydrogenases are released from these mitochondria by sonication, and these enzymes were partially purified using fast protein liquid chromatography. One of the enzymes, designated peak I, is capable of oxidizing NADPH and the β form of NADH. The other two activities, peaks II and III, oxidize only β-NADH. All three peaks are insensitive to divalent cation chelators and a complex I inhibitor, rotenone. The major component of peak I is a polypeptide with an apparent molecular mass of approximately 42 kilodaltons. Peak I activity was insensitive to platanetin, a specific inhibitor of the exogenous dehydrogenase, and insensitive to added Ca2+ or Mg2+. Peak I displayed a broad pH activity profile with an optimum between 7.5 and 8.0 for both NADPH and NADH. Purified peak II gave a single polypeptide of about 32 kilodaltons, had a pH optimum between 7.0 and 7.5, and was slightly stimulated by Ca2+ and Mg2+. As with peak I, platanetin had no effect on peak II activity. Peak III was not purified completely, but contained two major polypeptides with apparent molecular masses of 55 and 40 kilodaltons. This enzyme was not affected by Ca2+ and Mg2+, but was inhibited by platanetin. The peak III enzyme had a rather sharp pH optimum of approximately 6.5 to 6.6. The above data indicate that peak III activity is likely the exogenous NADH dehydrogenase.

Original languageEnglish (US)
Pages (from-to)1317-1322
Number of pages6
JournalPlant Physiology
Volume97
Issue number4
DOIs
StatePublished - Jan 1 1991

Fingerprint

Beta vulgaris
NAD(P)H dehydrogenase (quinone)
NADH dehydrogenase (ubiquinone)
NAD
NAD (coenzyme)
Mitochondria
Oxidoreductases
mitochondria
NADH Dehydrogenase
polypeptides
Enzymes
enzymes
NADP
calcium
NADP (coenzyme)
Peptides
molecular weight
Rotenone
rotenone
Sonication

ASJC Scopus subject areas

  • Physiology
  • Genetics
  • Plant Science

Cite this

Partial purification and characterization of three NAD(P)H dehydrogenases from Beta vulgaris mitochondria. / Luethy, Michael H.; Hayes, Marianne K.; Elthon, Thomas E.

In: Plant Physiology, Vol. 97, No. 4, 01.01.1991, p. 1317-1322.

Research output: Contribution to journalArticle

Luethy, Michael H. ; Hayes, Marianne K. ; Elthon, Thomas E. / Partial purification and characterization of three NAD(P)H dehydrogenases from Beta vulgaris mitochondria. In: Plant Physiology. 1991 ; Vol. 97, No. 4. pp. 1317-1322.
@article{1fe81a5bd4d04851a59147eb554be7f3,
title = "Partial purification and characterization of three NAD(P)H dehydrogenases from Beta vulgaris mitochondria",
abstract = "Mitochondria isolated from the taproot of beet (Beta vulgaris) were used in an effort to identify and partially purify the proteins constituting the exogenous NADH dehydrogenase. Three NAD(P)H dehydrogenases are released from these mitochondria by sonication, and these enzymes were partially purified using fast protein liquid chromatography. One of the enzymes, designated peak I, is capable of oxidizing NADPH and the β form of NADH. The other two activities, peaks II and III, oxidize only β-NADH. All three peaks are insensitive to divalent cation chelators and a complex I inhibitor, rotenone. The major component of peak I is a polypeptide with an apparent molecular mass of approximately 42 kilodaltons. Peak I activity was insensitive to platanetin, a specific inhibitor of the exogenous dehydrogenase, and insensitive to added Ca2+ or Mg2+. Peak I displayed a broad pH activity profile with an optimum between 7.5 and 8.0 for both NADPH and NADH. Purified peak II gave a single polypeptide of about 32 kilodaltons, had a pH optimum between 7.0 and 7.5, and was slightly stimulated by Ca2+ and Mg2+. As with peak I, platanetin had no effect on peak II activity. Peak III was not purified completely, but contained two major polypeptides with apparent molecular masses of 55 and 40 kilodaltons. This enzyme was not affected by Ca2+ and Mg2+, but was inhibited by platanetin. The peak III enzyme had a rather sharp pH optimum of approximately 6.5 to 6.6. The above data indicate that peak III activity is likely the exogenous NADH dehydrogenase.",
author = "Luethy, {Michael H.} and Hayes, {Marianne K.} and Elthon, {Thomas E}",
year = "1991",
month = "1",
day = "1",
doi = "10.1104/pp.97.4.1317",
language = "English (US)",
volume = "97",
pages = "1317--1322",
journal = "Plant Physiology",
issn = "0032-0889",
publisher = "American Society of Plant Biologists",
number = "4",

}

TY - JOUR

T1 - Partial purification and characterization of three NAD(P)H dehydrogenases from Beta vulgaris mitochondria

AU - Luethy, Michael H.

AU - Hayes, Marianne K.

AU - Elthon, Thomas E

PY - 1991/1/1

Y1 - 1991/1/1

N2 - Mitochondria isolated from the taproot of beet (Beta vulgaris) were used in an effort to identify and partially purify the proteins constituting the exogenous NADH dehydrogenase. Three NAD(P)H dehydrogenases are released from these mitochondria by sonication, and these enzymes were partially purified using fast protein liquid chromatography. One of the enzymes, designated peak I, is capable of oxidizing NADPH and the β form of NADH. The other two activities, peaks II and III, oxidize only β-NADH. All three peaks are insensitive to divalent cation chelators and a complex I inhibitor, rotenone. The major component of peak I is a polypeptide with an apparent molecular mass of approximately 42 kilodaltons. Peak I activity was insensitive to platanetin, a specific inhibitor of the exogenous dehydrogenase, and insensitive to added Ca2+ or Mg2+. Peak I displayed a broad pH activity profile with an optimum between 7.5 and 8.0 for both NADPH and NADH. Purified peak II gave a single polypeptide of about 32 kilodaltons, had a pH optimum between 7.0 and 7.5, and was slightly stimulated by Ca2+ and Mg2+. As with peak I, platanetin had no effect on peak II activity. Peak III was not purified completely, but contained two major polypeptides with apparent molecular masses of 55 and 40 kilodaltons. This enzyme was not affected by Ca2+ and Mg2+, but was inhibited by platanetin. The peak III enzyme had a rather sharp pH optimum of approximately 6.5 to 6.6. The above data indicate that peak III activity is likely the exogenous NADH dehydrogenase.

AB - Mitochondria isolated from the taproot of beet (Beta vulgaris) were used in an effort to identify and partially purify the proteins constituting the exogenous NADH dehydrogenase. Three NAD(P)H dehydrogenases are released from these mitochondria by sonication, and these enzymes were partially purified using fast protein liquid chromatography. One of the enzymes, designated peak I, is capable of oxidizing NADPH and the β form of NADH. The other two activities, peaks II and III, oxidize only β-NADH. All three peaks are insensitive to divalent cation chelators and a complex I inhibitor, rotenone. The major component of peak I is a polypeptide with an apparent molecular mass of approximately 42 kilodaltons. Peak I activity was insensitive to platanetin, a specific inhibitor of the exogenous dehydrogenase, and insensitive to added Ca2+ or Mg2+. Peak I displayed a broad pH activity profile with an optimum between 7.5 and 8.0 for both NADPH and NADH. Purified peak II gave a single polypeptide of about 32 kilodaltons, had a pH optimum between 7.0 and 7.5, and was slightly stimulated by Ca2+ and Mg2+. As with peak I, platanetin had no effect on peak II activity. Peak III was not purified completely, but contained two major polypeptides with apparent molecular masses of 55 and 40 kilodaltons. This enzyme was not affected by Ca2+ and Mg2+, but was inhibited by platanetin. The peak III enzyme had a rather sharp pH optimum of approximately 6.5 to 6.6. The above data indicate that peak III activity is likely the exogenous NADH dehydrogenase.

UR - http://www.scopus.com/inward/record.url?scp=0000813770&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=0000813770&partnerID=8YFLogxK

U2 - 10.1104/pp.97.4.1317

DO - 10.1104/pp.97.4.1317

M3 - Article

VL - 97

SP - 1317

EP - 1322

JO - Plant Physiology

JF - Plant Physiology

SN - 0032-0889

IS - 4

ER -