1V16

CROSSTALK BETWEEN COFACTOR BINDING AND THE PHOSPHORYLATION LOOP CONFORMATION IN THE BCKD MACHINE


Experimental Data Snapshot

  • Method: X-RAY DIFFRACTION
  • Resolution: 1.90 Å
  • R-Value Free: 0.170 
  • R-Value Work: 0.131 
  • R-Value Observed: 0.132 

wwPDB Validation   3D Report Full Report


Ligand Structure Quality Assessment 


This is version 2.1 of the entry. See complete history


Literature

Cross-Talk between Thiamin Diphosphate Binding and Phosphorylation Loop Conformation in Human Branched-Chain {Alpha}-Keto Acid Decarboxylase/Dehydrogenase

Li, J.Wynn, R.M.Machius, M.Chuang, J.L.Karthikeyan, S.Tomchick, D.R.Chuang, D.T.

(2004) J Biol Chem 279: 32968

  • DOI: https://doi.org/10.1074/jbc.M403611200
  • Primary Citation of Related Structures:  
    1V11, 1V16, 1V1M, 1V1R

  • PubMed Abstract: 

    The decarboxylase/dehydrogenase (E1b) component of the 4-megadalton human branched-chain alpha-keto acid dehydrogenase (BCKD) metabolic machine is a thiamin diphosphate (ThDP)-dependent enzyme with a heterotetrameric cofactor-binding fold. The E1b component catalyzes the decarboxylation of alpha-keto acids and the subsequent reductive acylation of the lipoic acid-bearing domain (LBD) from the 24-meric transacylase (E2b) core. In the present study, we show that the binding of cofactor ThDP to the E1b active site induces a disorder-to-order transition of the conserved phosphorylation loop carrying the two phosphorylation sites Ser(292)-alpha and Ser(302)-alpha, as deduced from the 1.80-1.85 A apoE1b and holoE1b structures. The induced loop conformation is essential for the recognition of lipoylated LBD to initiate E1b-catalyzed reductive acylation. Alterations of invariant Arg(287)-alpha, Asp(295)-alpha, Tyr(300)-alpha, and Arg(301)-alpha that form a hydrogen-bonding network in the phosphorylation loop result in the disordering of the loop conformation as elucidated by limited proteolysis, accompanied by the impaired binding and diminished reductive acylation of lipoylated LBD. In contrast, k(cat) values for E1b-catalyzed decarboxylation of the alpha-keto acid are higher in these E1b mutants than in wild-type E1b, with higher K(m) values for the substrate in the mutants. ThDP binding that orders the loop prevents phosphorylation of E1b by the BCKD kinase and averts the inactivation of wild-type E1b, but not the above mutants, by this covalent modification. Our results establish that the cross-talk between the bound ThDP and the phosphorylation loop conformation serves as a feed-forward switch for multiple reaction steps in the BCKD metabolic machine.


  • Organizational Affiliation

    Department of Biochemistry, University of Texas Southwestern Medical Center, Dallas, Texas 75390-9038, USA.


Macromolecules
Find similar proteins by:  (by identity cutoff)  |  3D Structure
Entity ID: 1
MoleculeChains Sequence LengthOrganismDetailsImage
2-OXOISOVALERATE DEHYDROGENASE ALPHA SUBUNIT400Homo sapiensMutation(s): 1 
EC: 1.2.4.4
UniProt & NIH Common Fund Data Resources
Find proteins for P12694 (Homo sapiens)
Explore P12694 
Go to UniProtKB:  P12694
PHAROS:  P12694
GTEx:  ENSG00000248098 
Entity Groups  
Sequence Clusters30% Identity50% Identity70% Identity90% Identity95% Identity100% Identity
UniProt GroupP12694
Sequence Annotations
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  • Reference Sequence
Find similar proteins by:  (by identity cutoff)  |  3D Structure
Entity ID: 2
MoleculeChains Sequence LengthOrganismDetailsImage
2-OXOISOVALERATE DEHYDROGENASE BETA SUBUNIT342Homo sapiensMutation(s): 0 
EC: 1.2.4.4
UniProt & NIH Common Fund Data Resources
Find proteins for P21953 (Homo sapiens)
Explore P21953 
Go to UniProtKB:  P21953
PHAROS:  P21953
GTEx:  ENSG00000083123 
Entity Groups  
Sequence Clusters30% Identity50% Identity70% Identity90% Identity95% Identity100% Identity
UniProt GroupP21953
Sequence Annotations
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  • Reference Sequence
Small Molecules
Ligands 6 Unique
IDChains Name / Formula / InChI Key2D Diagram3D Interactions
TPP
Query on TPP

Download Ideal Coordinates CCD File 
I [auth A]THIAMINE DIPHOSPHATE
C12 H19 N4 O7 P2 S
AYEKOFBPNLCAJY-UHFFFAOYSA-O
BEN
Query on BEN

Download Ideal Coordinates CCD File 
J [auth A]BENZAMIDINE
C7 H8 N2
PXXJHWLDUBFPOL-UHFFFAOYSA-N
GOL
Query on GOL

Download Ideal Coordinates CCD File 
L [auth B],
M [auth B],
N [auth B]
GLYCEROL
C3 H8 O3
PEDCQBHIVMGVHV-UHFFFAOYSA-N
MN
Query on MN

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D [auth A]MANGANESE (II) ION
Mn
WAEMQWOKJMHJLA-UHFFFAOYSA-N
K
Query on K

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C [auth A],
K [auth B]
POTASSIUM ION
K
NPYPAHLBTDXSSS-UHFFFAOYSA-N
CL
Query on CL

Download Ideal Coordinates CCD File 
E [auth A],
F [auth A],
G [auth A],
H [auth A]
CHLORIDE ION
Cl
VEXZGXHMUGYJMC-UHFFFAOYSA-M
Experimental Data & Validation

Experimental Data

  • Method: X-RAY DIFFRACTION
  • Resolution: 1.90 Å
  • R-Value Free: 0.170 
  • R-Value Work: 0.131 
  • R-Value Observed: 0.132 
  • Space Group: P 31 2 1
Unit Cell:
Length ( Å )Angle ( ˚ )
a = 145.397α = 90
b = 145.397β = 90
c = 69.104γ = 120
Software Package:
Software NamePurpose
REFMACrefinement
HKL-2000data reduction
HKL-2000data scaling

Structure Validation

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Ligand Structure Quality Assessment 


Entry History 

Deposition Data

Revision History  (Full details and data files)

  • Version 1.0: 2004-06-03
    Type: Initial release
  • Version 1.1: 2011-07-13
    Changes: Advisory, Version format compliance
  • Version 1.2: 2019-05-15
    Changes: Data collection, Experimental preparation, Other
  • Version 1.3: 2019-09-25
    Changes: Data collection, Experimental preparation, Other
  • Version 2.0: 2021-08-04
    Changes: Atomic model, Data collection, Derived calculations, Non-polymer description, Structure summary
  • Version 2.1: 2023-12-13
    Changes: Data collection, Database references, Refinement description