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J. Biol. Chem., Vol. 277, Issue 40, 36905-36908, October 4, 2002
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-Keto Acid Dehydrogenase Kinase*
From the Departments of Biochemistry and Internal Medicine,
University of Texas Southwestern Medical Center, Dallas, Texas
75390
The branched-chain 
-keto acid dehydrogenase
(BCKD) kinase (abbreviated as BCK) down-regulates activity of the
mammalian mitochondrial BCKD complex by reversible phosphorylation of
the decarboxylase (E1b) component of the complex. The binding of BCK to
the holotransacylase (E2b) core of the BCKD complex results in the
stimulation of BCK activity. Here we show that the lipoylated lipoic
acid-bearing domain (lip-LBD) (residues 1-84) of E2b alone does not
interact with BCK. However, lip-LBD constructs containing various
lengths of the C-terminal hinge region of LBD are able to bind to BCK as measured by a newly developed solubility-based binding assay. Isothermal titration calorimetry measurements produced a dissociation constant of 8.06 × 10
6 M and
binding enthalpy of -3.68 kcal/mol for the interaction of BCK with a
construct containing lip-LBD and the Glu-Glu-Asp-Xaa-Xaa-Glu sequence
of the C-terminal hinge region of LBD.
These thermodynamic parameters are similar to those obtained for
binding of BCK to a lipoylated di-domain construct, which harbors LBD,
the entire hinge region, and the downstream subunit-binding domain of
E2b. Our data establish that the C-terminal hinge region of LBD
containing the above negatively charged residues is essential for the
interaction between the lip-LBD construct and BCK.
To whom correspondence should be addressed: Dept. of Biochemistry,
University of Texas Southwestern Medical Center, 5323 Harry Hines
Blvd., Dallas, TX 75390-9038. Tel.: 214-648-2457; Fax: 214-648-8856; E-mail: David.Chuang@UTSouthwestern.edu.
This article has been cited by other articles:
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  C.-F. Chang, H.-T. Chou, Y.-J. Lin, S.-J. Lee, J. L. Chuang, D. T. Chuang, and T.-h. Huang Structure of the Subunit Binding Domain and Dynamics of the Di-domain Region from the Core of Human Branched Chain {alpha}-Ketoacid Dehydrogenase Complex J. Biol. Chem., September 22, 2006; 281(38): 28345 - 28353. [Abstract] [Full Text] [PDF]
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J. Li, R. M. Wynn, M. Machius, J. L. Chuang, S. Karthikeyan, D. R. Tomchick, and D. T. Chuang Cross-talk between Thiamin Diphosphate Binding and Phosphorylation Loop Conformation in Human Branched-chain {alpha}-Keto Acid Decarboxylase/Dehydrogenase J. Biol. Chem., July 30, 2004; 279(31): 32968 - 32978. [Abstract] [Full Text] [PDF]
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J. L. Chuang, R. M. Wynn, C. C. Moss, J.-l. Song, J. Li, N. Awad, H. Mandel, and D. T. Chuang Structural and Biochemical Basis for Novel Mutations in Homozygous Israeli Maple Syrup Urine Disease Patients: A PROPOSED MECHANISM FOR THE THIAMIN-RESPONSIVE PHENOTYPE J. Biol. Chem., April 23, 2004; 279(17): 17792 - 17800. [Abstract] [Full Text] [PDF]
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R. M. Wynn, M. Machius, J. L. Chuang, J. Li, D. R. Tomchick, and D. T. Chuang Roles of His291-{alpha} and His146-{beta}' in the Reductive Acylation Reaction Catalyzed by Human Branched-chain {alpha}-Ketoacid Dehydrogenase: REFINED PHOSPHORYLATION LOOP STRUCTURE IN THE ACTIVE SITE J. Biol. Chem., October 31, 2003; 278(44): 43402 - 43410. [Abstract] [Full Text] [PDF]
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