PDBsum entry 1c0p

Oxidoreductase

PDB id: 1c0p

Contents

Protein chain

363 a.a. *

Ligands

FAD-DAL-PER

GOL

Waters ×612

* Residue conservation analysis

PDB id:

1c0p

Name:

Oxidoreductase

Title:

D-amino acic oxidase in complex with d-alanine and a partially occupied biatomic species

Structure:

D-amino acid oxidase. Chain: a. Engineered: yes

Source:

Rhodosporidium toruloides. Organism_taxid: 5286. Expressed in: escherichia coli. Expression_system_taxid: 562.

Biol. unit:

Dimer (from PDB file)

Resolution:

1.20Å R-factor: 0.125 R-free: 0.150

Authors:

S.Umhau,L.Pollegioni,G.Molla,K.Diederichs,W.Welte,S.M.Pilone,S.Ghisla

Key ref:

S.Umhau et al. (2000). The x-ray structure of D-amino acid oxidase at very high resolution identifies the chemical mechanism of flavin-dependent substrate dehydrogenation. Proc Natl Acad Sci U S A, 97, 12463-12468. PubMed id: 11070076 DOI: 10.1073/pnas.97.23.12463

Date:

19-Jul-99 Release date: 22-Nov-00

Protein chain

P80324  (OXDA_RHOTO) -  D-amino-acid oxidase from Rhodotorula toruloides

Seq: 368 a.a.

Struc: 363 a.a.

Enzyme reactions

• Enzyme class: E.C.1.4.3.3 - D-amino-acid oxidase.
• Pathway: Cephalosporin Biosynthesis
• Reaction: a D-alpha-amino acid + O2 + H2O = a 2-oxocarboxylate + H2O2 + NH4⁺

D-alpha-amino acid + O2 + H2O (Bound ligand (Het Group name = PER) corresponds exactly) = 2-oxocarboxylate (Bound ligand (Het Group name = GOL) matches with 71.43% similarity) + H2O2 + NH4(+)

• Cofactor: FAD

FAD (Bound ligand (Het Group name = FAD) corresponds exactly)

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

reference

DOI no: 10.1073/pnas.97.23.12463

Proc Natl Acad Sci U S A 97:12463-12468 (2000)

PubMed id: 11070076

The x-ray structure of D-amino acid oxidase at very high resolution identifies the chemical mechanism of flavin-dependent substrate dehydrogenation.

S.Umhau, L.Pollegioni, G.Molla, K.Diederichs, W.Welte, M.S.Pilone, S.Ghisla.

ABSTRACT

Flavin is one of the most versatile redox cofactors in nature and is used by many enzymes to perform a multitude of chemical reactions. d-Amino acid oxidase (DAAO), a member of the flavoprotein oxidase family, is regarded as a key enzyme for the understanding of the mechanism underlying flavin catalysis. The very high-resolution structures of yeast DAAO complexed with d-alanine, d-trifluoroalanine, and l-lactate (1.20, 1.47, and 1.72 A) provide strong evidence for hydride transfer as the mechanism of dehydrogenation. This is inconsistent with the alternative carbanion mechanism originally favored for this type of enzymatic reaction. The step of hydride transfer can proceed without involvement of amino acid functional groups. These structures, together with results from site-directed mutagenesis, point to orbital orientation/steering as the major factor in catalysis. A diatomic species, proposed to be a peroxide, is found at the active center and on the Re-side of the flavin. These results are of general relevance for the mechanisms of flavoproteins and lead to the proposal of a common dehydrogenation mechanism for oxidases and dehydrogenases.

Selected figure(s)

Figure 3.
Fig. 3. Active site of RgDAAO at 1.2-Å resolution. Stereo view of the 2F[obs] F[calc] map (orange, 3 ) and the omit map (magenta, 3 ) showing clear electron density assigned to a peroxide species. The data were obtained from RgDAAO crystals soaked with 20 mM D-alanine and 200 mM pyruvate.

Figure 4.
Fig. 4. Reciprocal orientation of ligands and flavin plane. For clarity, the dioxygen species has been omitted. Dashed lines represent H-bonds. (A) D-Ala is viewed along its N- C axis; the electron density is shown at 2 . (B) D-F[3]-Ala (Upper) and L-lactate (Lower). The green trace represents the ideal line connecting the flavin N(5) and the ligand C centers (distance 3.2 Å). Note that the C---H function (grey) of L-lactate points away from the flavin. With L-lactate, the position of the ---H results from H-inclusion in the refinement. In the case of D-F[3]-Ala (A), the number of observations (1.72 Å) does not allow positioning. The strong H-bond interactions with the -NH[2]/OH, together with the electrostatic interaction of the substrate carboxylate group with Arg-285, Tyr-238, and Tyr-223, provide the rationale for substrate D-specificity in that they prevent binding of the L-amino acid in a productive manner.

Figures were selected by the author.