PDBsum entry 1j9z

Oxidoreductase

PDB id: 1j9z

Contents

Protein chains

613 a.a. *

Ligands

FAD ×2

FMN ×2

NAP ×2

Waters ×210

* Residue conservation analysis

PDB id:

1j9z

Name:

Oxidoreductase

Title:

Cypor-w677g

Structure:

NADPH-cytochrome p450 reductase. Chain: a, b. Synonym: NADPH-cytochrome p450 oxidoreductase. Engineered: yes. Mutation: yes

Source:

Rattus norvegicus. Norway rat. Organism_taxid: 10116. Gene: cypor. Expressed in: escherichia coli. Expression_system_taxid: 562.

Resolution:

2.70Å R-factor: 0.188 R-free: 0.270

Authors:

P.A.Hubbard,A.L.Shen,R.Paschke,C.B.Kasper,J.J.Kim

Key ref:

P.A.Hubbard et al. (2001). NADPH-cytochrome P450 oxidoreductase. Structural basis for hydride and electron transfer. J Biol Chem, 276, 29163-29170. PubMed id: 11371558 DOI: 10.1074/jbc.M101731200

Date:

29-May-01 Release date: 22-Aug-01

Protein chains

P00388  (NCPR_RAT) -  NADPH--cytochrome P450 reductase from Rattus norvegicus

Seq: 678 a.a.

Struc: 613 a.a.*

* PDB and UniProt seqs differ at 1 residue position (black cross)

Enzyme reactions

• Enzyme class: E.C.1.6.2.4 - NADPH--hemoprotein reductase.
• Reaction: 2 oxidized [cytochrome P450] + NADPH = 2 reduced [cytochrome P450] + NADP⁺ + H⁺

2 × oxidized [cytochrome P450] (Bound ligand (Het Group name = NAP) corresponds exactly) + NADPH = 2 × reduced [cytochrome P450] + NADP(+) + H(+)

• Cofactor: FAD; FMN

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

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

reference

DOI no: 10.1074/jbc.M101731200

J Biol Chem 276:29163-29170 (2001)

PubMed id: 11371558

NADPH-cytochrome P450 oxidoreductase. Structural basis for hydride and electron transfer.

P.A.Hubbard, A.L.Shen, R.Paschke, C.B.Kasper, J.J.Kim.

ABSTRACT

NADPH-cytochrome P450 oxidoreductase catalyzes transfer of electrons from NADPH, via two flavin cofactors, to various cytochrome P450s. The crystal structure of the rat reductase complexed with NADP(+) has revealed that nicotinamide access to FAD is blocked by an aromatic residue (Trp-677), which stacks against the re-face of the isoalloxazine ring of the flavin. To investigate the nature of interactions between the nicotinamide, FAD, and Trp-677 during the catalytic cycle, three mutant proteins were studied by crystallography. The first mutant, W677X, has the last two C-terminal residues, Trp-677 and Ser-678, removed; the second mutant, W677G, retains the C-terminal serine residue. The third mutant has the following three catalytic residues substituted: S457A, C630A, and D675N. In the W677X and W677G structures, the nicotinamide moiety of NADP(+) lies against the FAD isoalloxazine ring with a tilt of approximately 30 degrees between the planes of the two rings. These results, together with the S457A/C630A/D675N structure, allow us to propose a mechanism for hydride transfer regulated by changes in hydrogen bonding and pi-pi interactions between the isoalloxazine ring and either the nicotinamide ring or Trp-677 indole ring. Superimposition of the mutant and wild-type structures shows significant mobility between the two flavin domains of the enzyme. This, together with the high degree of disorder observed in the FMN domain of all three mutant structures, suggests that conformational changes occur during catalysis.

Selected figure(s)

Figure 2.
Fig. 2. Stereo diagram of an F[o] F[c] OMIT map contoured at 2 from the W667X deletion mutant. The density shows the A-face of the nicotinamide ring of NADP+ to stack against the re-face of the isoalloxazine ring of FAD, with a ~30° tilt between the planes of the two rings.

Figure 4.
Fig. 4. Diagram showing the hydrogen bond networks surrounding the isoalloxazine ring of FAD and the nicotinamide ring of NADP+. Thin dashed lines represent hydrogen bonds; thick dotted lines are van der Waal's contacts. A, the wild-type structure. The two catalytic waters involved in hydrogen bond networks with FAD are included. B, the W677X mutant structure. The presence of the nicotinamide ring results in Asp-675 moving out of hydrogen bonding distance from Ser-457 and Cys-630. Van der Waal's contacts are formed between the C-4 atom of the nicotinamide ring and Cys-630 and from Ser-457 to the N-5 atom of FAD. Another van der Waal's contact is formed between the C-4 atom of the nicotinamide ring and the N-5 atom of FAD.

The above figures are reprinted by permission from the ASBMB: J Biol Chem (2001, 276, 29163-29170) copyright 2001.

Figures were selected by an automated process.