PDBsum entry: 2ozl

Oxidoreductase

PDB id: 2ozl

Contents

Protein chains

362 a.a. *

330 a.a. *

Ligands

TPP ×2

Metals

__K ×2

_MG ×2

Waters ×757

* Residue conservation analysis

PDB id:

2ozl

Name:

Oxidoreductase

Title:

Human pyruvate dehydrogenase s264e variant

Structure:

Pyruvate dehydrogenase e1 component alpha subunit, somatic form. Chain: a, c. Fragment: alpha subunit. Synonym: pdhe1-a type i. Engineered: yes. Mutation: yes. Pyruvate dehydrogenase e1 component subunit beta. Chain: b, d.

Source:

Homo sapiens. Human. Organism_taxid: 9606. Gene: pdha1, phe1a. Expressed in: escherichia coli. Expression_system_taxid: 562. Gene: pdhb, phe1b.

Resolution:

1.90Å R-factor: 0.186 R-free: 0.221

Authors:

E.M.Ciszak,P.M.Dominiak,M.S.Patel,L.G.Korotchkina

Key ref:

F.Seifert et al. (2007). Phosphorylation of serine 264 impedes active site accessibility in the E1 component of the human pyruvate dehydrogenase multienzyme complex. Biochemistry, 46, 6277-6287. PubMed id: 17474719 DOI: 10.1021/bi700083z

Date:

26-Feb-07 Release date: 22-May-07

Protein chains

P08559  (ODPA_HUMAN) -  Pyruvate dehydrogenase E1 component subunit alpha, somatic form, mitochondrial

Seq: 390 a.a.

Struc: 362 a.a.*

Protein chains

P11177  (ODPB_HUMAN) -  Pyruvate dehydrogenase E1 component subunit beta, mitochondrial

Seq: 359 a.a.

Struc: 330 a.a.*

* PDB and UniProt seqs differ at 3 residue positions (black crosses)

Enzyme reactions

• Enzyme class: Chains A, B, C, D: E.C.1.2.4.1 - Pyruvate dehydrogenase (acetyl-transferring).
• Pathway: Oxo-acid dehydrogenase complexes
• Reaction: Pyruvate + [dihydrolipoyllysine-residue acetyltransferase] lipoyllysine = [dihydrolipoyllysine-residue acetyltransferase] S-acetyldihydrolipoyllysine + CO₂
• Cofactor: Thiamine diphosphate

Thiamine diphosphate (Bound ligand (Het Group name = TPP) corresponds exactly)

Molecule diagrams generated from .mol files obtained from the KEGG ftp site

 Gene Ontology (GO) functional annotation 

• Cellular component: intracellular membrane-bounded organelle (4 terms)
• Biological process: metabolic process (6 terms)
• Biochemical function: catalytic activity (5 terms)

reference

DOI no: 10.1021/bi700083z

Biochemistry 46:6277-6287 (2007)

PubMed id: 17474719

Phosphorylation of serine 264 impedes active site accessibility in the E1 component of the human pyruvate dehydrogenase multienzyme complex.

F.Seifert, E.Ciszak, L.Korotchkina, R.Golbik, M.Spinka, P.Dominiak, S.Sidhu, J.Brauer, M.S.Patel, K.Tittmann.

ABSTRACT

At the junction of glycolysis and the Krebs cycle in cellular metabolism, the pyruvate dehydrogenase multienzyme complex (PDHc) catalyzes the oxidative decarboxylation of pyruvate to acetyl-CoA. In mammals, PDHc is tightly regulated by phosphorylation-dephosphorylation of three serine residues in the thiamin-dependent pyruvate dehydrogenase (E1) component. In vivo, inactivation of human PDHc correlates mostly with phosphorylation of serine 264, which is located at the entrance of the substrate channel leading to the active site of E1. Despite intense investigations, the molecular mechanism of this inactivation has remained enigmatic. Here, a detailed analysis of microscopic steps of catalysis in human wild-type PDHc-E1 and pseudophosphorylation variant Ser264Glu elucidates how phosphorylation of Ser264 affects catalysis. Whereas the intrinsic reactivity of the active site in catalysis of pyruvate decarboxylation remains nearly unaltered, the preceding binding of substrate to the enzyme's active site via the substrate channel and the subsequent reductive acetylation of the E2 component are severely slowed in the phosphorylation variant. The structure of pseudophosphorylation variant Ser264Glu determined by X-ray crystallography reveals no differences in the three-dimensional architecture of the phosphorylation loop or of the active site, when compared to those of the wild-type enzyme. However, the channel leading to the active site is partially obstructed by the side chain of residue 264 in the variant. By analogy, a similar obstruction of the substrate channel can be anticipated to result from a phosphorylation of Ser264. The kinetic and thermodynamic results in conjunction with the structure of Ser264Glu suggest that phosphorylation blocks access to the active site by imposing a steric and electrostatic barrier for substrate binding and active site coupling with the E2 component. As a Ser264Gln variant, which carries no charge at position 264, is also selectively deficient in pyruvate binding and reductive acetylation of E2, we conclude that mostly steric effects account for inhibition of PDHc by phosphorylation.