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

This Article Full Text Full Text (PDF) 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 Davoodi, J. Articles by Hutson, S. M. Search for Related Content PubMed PubMed Citation Articles by Davoodi, J. Articles by Hutson, S. M. Social Bookmarking                      What's this?

J Biol Chem, Vol. 273, Issue 9, 4982-4989, February 27, 1998

Overexpression and Characterization of the Human Mitochondrial and Cytosolic Branched-chain Aminotransferases

Jamshid Davoodi, Penny M. Drown, Randy K. Bledsoe^§, Reidar Wallin^¶, Gregory D. Reinhart, and Susan M. Hutson

From the Departments of  Biochemistry and ^¶ Internal Medicine, Wake Forest University School of Medicine, Winston Salem, North Carolina 27157, ^§ Glaxo Wellcome Research and Development, Research Triangle Park, North Carolina 27709, and the  Department of Biochemistry and Biophysics, Texas A & M University, College Station, Texas 77843-2128

We have developed overexpression systems for the human branched-chain aminotransferase isoenzymes. The enzymes function as dimers and have substrate specificity comparable with the rat enzymes. The human cytosolic enzyme appears to turn over 2-5 times faster than the mitochondrial enzyme, and there may be anion and cation effects on the kinetics of both enzymes. The two proteins demonstrate similar absorption profiles, and the far UV circular dichroism spectra show that no global structural changes occur when the proteins are converted from the pyridoxal to pyridoxamine form. On the other hand, the near UV circular dichroism spectra suggest differences in the local environment surrounding tyrosines within these proteins. Both enzymes require a reducing environment for maximal activity, but the mitochondrial enzyme can be inhibited by nickel ions in the presence of reducing agents, while the cytosolic enzyme is unaffected. Chemical denaturation profiles of the proteins show that there are differences in structural stability. Titration of -SH groups with 5,5'-dithiobis(2-nitrobenzoic acid) suggests that no disulfide bonds are present in the mitochondrial enzyme and that at least two disulfide bonds are present in the cytosolic enzyme. Two -SH groups are titrated in the native form of the mitochondrial enzyme, leading to complete inhibition of activity, while only one -SH group is titrated in the cytosolic enzyme with no effect on activity. Although these proteins share 58% identity in primary amino acid sequence, the local environment surrounding the active site appears unique for each isoenzyme.

CiteULike

Complore

Connotea

Del.icio.us

Digg

Reddit

Technorati

This article has been cited by other articles:

				M. M. Islam, R. Wallin, R. M. Wynn, M. Conway, H. Fujii, J. A. Mobley, D. T. Chuang, and S. M. Hutson A Novel Branched-chain Amino Acid Metabolon: PROTEIN-PROTEIN INTERACTIONS IN A SUPRAMOLECULAR COMPLEX J. Biol. Chem., April 20, 2007; 282(16): 11893 - 11903. [Abstract] [Full Text] [PDF]

				N. H. Yennawar, M. M. Islam, M. Conway, R. Wallin, and S. M. Hutson Human Mitochondrial Branched Chain Aminotransferase Isozyme: STRUCTURAL ROLE OF THE CXXC CENTER IN CATALYSIS J. Biol. Chem., December 22, 2006; 281(51): 39660 - 39671. [Abstract] [Full Text] [PDF]

				M. Goto, I. Miyahara, K. Hirotsu, M. Conway, N. Yennawar, M. M. Islam, and S. M. Hutson Structural Determinants for Branched-chain Aminotransferase Isozyme-specific Inhibition by the Anticonvulsant Drug Gabapentin J. Biol. Chem., November 4, 2005; 280(44): 37246 - 37256. [Abstract] [Full Text] [PDF]

				M. E. Rabaglia, M. P. Gray-Keller, B. L. Frey, M. R. Shortreed, L. M. Smith, and A. D. Attie {alpha}-Ketoisocaproate-induced hypersecretion of insulin by islets from diabetes-susceptible mice Am J Physiol Endocrinol Metab, August 1, 2005; 289(2): E218 - E224. [Abstract] [Full Text] [PDF]

				S. M. Hutson, A. J. Sweatt, and K. F. LaNoue Branched-Chain Amino Acid Metabolism: Implications for Establishing Safe Intakes J. Nutr., June 1, 2005; 135(6): 1557S - 1564S. [Abstract] [Full Text] [PDF]

				N. Wajih, D. C. Sane, S. M. Hutson, and R. Wallin Engineering of a Recombinant Vitamin K-dependent {gamma}-Carboxylation System with Enhanced {gamma}-Carboxyglutamic Acid Forming Capacity: EVIDENCE FOR A FUNCTIONAL CXXC REDOX CENTER IN THE SYSTEM J. Biol. Chem., March 18, 2005; 280(11): 10540 - 10547. [Abstract] [Full Text] [PDF]

				J. J. Vermeersch, S. Christmann-Franck, L. V. Karabashyan, S. Fermandjian, G. Mirambeau, and P. A. Der Garabedian Pyridoxal 5'-phosphate inactivates DNA topoisomerase IB by modifying the lysine general acid Nucleic Acids Res., October 19, 2004; 32(18): 5649 - 5657. [Abstract] [Full Text] [PDF]

				N. Wajih, D. C. Sane, S. M. Hutson, and R. Wallin The Inhibitory Effect of Calumenin on the Vitamin K-dependent {gamma}-Carboxylation System: CHARACTERIZATION OF THE SYSTEM IN NORMAL AND WARFARIN-RESISTANT RATS J. Biol. Chem., June 11, 2004; 279(24): 25276 - 25283. [Abstract] [Full Text] [PDF]

				A. J. Sweatt, M. Wood, A. Suryawan, R. Wallin, M. C. Willingham, and S. M. Hutson Branched-chain amino acid catabolism: unique segregation of pathway enzymes in organ systems and peripheral nerves Am J Physiol Endocrinol Metab, January 1, 2004; 286(1): E64 - E76. [Abstract] [Full Text]

				G. F. Sanchez-Perez, M. Gasset, J. J. Calvete, and M. A. Pajares Role of an Intrasubunit Disulfide in the Association State of the Cytosolic Homo-oligomer Methionine Adenosyltransferase J. Biol. Chem., February 21, 2003; 278(9): 7285 - 7293. [Abstract] [Full Text] [PDF]

				C. J. Lynch, S. M. Hutson, B. J. Patson, A. Vaval, and T. C. Vary Tissue-specific effects of chronic dietary leucine and norleucine supplementation on protein synthesis in rats Am J Physiol Endocrinol Metab, October 1, 2002; 283(4): E824 - E835. [Abstract] [Full Text] [PDF]

				R. Diebold, J. Schuster, K. Daschner, and S. Binder The Branched-Chain Amino Acid Transaminase Gene Family in Arabidopsis Encodes Plastid and Mitochondrial Proteins Plant Physiology, June 1, 2002; 129(2): 540 - 550. [Abstract] [Full Text] [PDF]

				L. C. Berger, J. Wilson, P. Wood, and B. J. Berger Methionine Regeneration and Aspartate Aminotransferase in Parasitic Protozoa J. Bacteriol., August 1, 2001; 183(15): 4421 - 4434. [Abstract] [Full Text] [PDF]

				M. G. Bixel, Y. Shimomura, S. M. Hutson, and B. Hamprecht Distribution of Key Enzymes of Branched-chain Amino Acid Metabolism in Glial and Neuronal Cells in Culture J. Histochem. Cytochem., March 1, 2001; 49(3): 407 - 418. [Abstract] [Full Text]

				A. R. Tovar, E. Becerril, R. Hernandez-Pando, G. Lopez, A. Suryawan, S. Desantiago, S. M. Hutson, and N. Torres Localization and expression of BCAT during pregnancy and lactation in the rat mammary gland Am J Physiol Endocrinol Metab, March 1, 2001; 280(3): E480 - E488. [Abstract] [Full Text] [PDF]

				G. Xu, G. Kwon, C. A. Marshall, T.-A. Lin, J. C. Lawrence Jr., and M. L. McDaniel Branched-chain Amino Acids Are Essential in the Regulation of PHAS-I and p70 S6 Kinase by Pancreatic beta -Cells. A POSSIBLE ROLE IN PROTEIN TRANSLATION AND MITOGENIC SIGNALING J. Biol. Chem., October 23, 1998; 273(43): 28178 - 28184. [Abstract] [Full Text] [PDF]