HOME HELP FEEDBACK SUBSCRIPTIONS ARCHIVE SEARCH TABLE OF CONTENTS		QUICK SEARCH:   [advanced] Author: Keyword(s): Year:  Vol:  Page:

J. Biol. Chem., Vol. 279, Issue 26, 27278-27285, June 25, 2004

This Article Full Text Full Text (PDF) All Versions of this Article: 279/26/27278    most recent M314148200v1 Submit a Letter to Editor Alert me when this article is cited Alert me when eLetters are posted Alert me if a correction is posted Citation Map Services Email this article to a friend Similar articles in this journal Similar articles in PubMed Alert me to new issues of the journal Download to citation manager Request Permissions Citing Articles Citing Articles via HighWire Citing Articles via Google Scholar Google Scholar Articles by MacDonald, M. J. Articles by Baker, T. R. Search for Related Content PubMed PubMed Citation Articles by MacDonald, M. J. Articles by Baker, T. R. Social Bookmarking                      What's this?

Immunochemical Identification of Coenzyme Q₀-Dihydrolipoamide Adducts in the E2 Components of the -Ketoglutarate and Pyruvate Dehydrogenase Complexes Partially Explains the Cellular Toxicity of Coenzyme Q₀^*

Michael J. MacDonald, Rhonda D. Husain¶, Susanne Hoffmann-Benning¶, and Tracie R. Baker

From the Childrens Diabetes Center, University of Wisconsin Medical School, Madison, Wisconsin 53706 and ^¶Mass Spectrometry Facility, Michigan State University, East Lansing, Michigan 48824

Coenzyme Q₀ (Q₀), a strong electrophile, is toxic to insulin-producing cells. Q₀ was incubated with rat and human pancreatic islets and INS-1 insulinoma cells, and its attachment to cellular proteins was studied with Western analysis using antiserum raised against the benzoquinone ring structure of ubiquinone (anti-Q). Q₀ covalently bonded to two proteins, one of 50 kDa and another of 70 kDa. Both proteins were found to be mitochondrial in human and rat islet cells and in many rat organs. Mitochondria were incubated with Q₀, and affinity-purified anti-Q was used to immunoprecipitate the 50-kDa protein. Amino acid sequencing identified it as dihydrolipoamide succinyltransferase, the E2 component of the -ketoglutarate dehydrogenase complex (KDC). Western analysis also showed that Q bonds to the E2 components of the purified KDC and ⁰the pyruvate dehydrogenase complex (PDC). Dihydrolipoamide acetyltransferase, the E2 of the PDC, has a molecular mass of 70 kDa, and the 70-kDa protein was inferred to be this enzyme. Q₀ was found to bond only to proteins containing dihydrolipoate, and in preparations of mitochondria, thiol reducing agents facilitated the attachment of Q₀, but oxidizing agents prevented it, suggesting that Q₀ bonds to thiols of dihydrolipoamide. Incubation of human or pig PDC with Q₀ followed by matrix-assisted laser desorption ionization time-of-flight and liquid chromatography/electrospray ionization mass spectrometry analyses of chymotrypsin-digested peptides of PDC E2 confirmed that Q₀ bonds to the dihydrolipoamide in these proteins. In mitochondria, coenzymes Q₁ and Q₂ did not bond to the 50-kDa protein but competed with the bonding of Q₀ to this protein. The prevention by Q₁ of characteristics the bonding of Q₀ to KDC E2, as well as other of the Q₀ effect, are reminiscent of the action of Q₀ on the mitochondrial permeability transition pore described previously (Fontaine, E., Ichas, F., and Bernardi, P. (1998) J. Biol. Chem. 273, 25734–25740).

Received for publication, December 24, 2003 , and in revised form, April 9, 2004.

^* This study was supported by National Institutes of Health Grant DK28348, the Oscar C. Rennebohm Foundation, and the Robert Wood Johnson Family Trust. The costs of publication of this article were defrayed in part by the payment of page charges. This article must therefore be hereby marked "advertisement" in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.

To whom correspondence should be addressed: Rm. 3459 Medical Science Center, 1300 University Ave., Madison, WI 53706. Tel.: 608-262-1195; Fax: 608-262-9300; E-mail: mjmacdon{at}wisc.edu.

CiteULike    Complore    Connotea    Del.icio.us    Digg    Reddit    Technorati    What's this?

This article has been cited by other articles:

				G. K. Scott, C. Atsriku, P. Kaminker, J. Held, B. Gibson, M. A. Baldwin, and C. C. Benz Vitamin K3 (Menadione)-Induced Oncosis Associated with Keratin 8 Phosphorylation and Histone H3 Arylation Mol. Pharmacol., September 1, 2005; 68(3): 606 - 615. [Abstract] [Full Text] [PDF]