PDBsum entry 1ja1

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Oxidoreductase PDB id
Protein chains
616 a.a. *
FAD ×2
FMN ×2
NAP ×2
Waters ×1126
* Residue conservation analysis
PDB id:
Name: Oxidoreductase
Title: Cypor-triple mutant
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.
1.80Å     R-factor:   0.208     R-free:   0.234
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
29-May-01     Release date:   22-Aug-01    
Go to PROCHECK summary

Protein chains
Pfam   ArchSchema ?
P00388  (NCPR_RAT) -  NADPH--cytochrome P450 reductase
678 a.a.
616 a.a.*
Key:    PfamA domain  Secondary structure  CATH domain
* PDB and UniProt seqs differ at 3 residue positions (black crosses)

 Enzyme reactions 
   Enzyme class: E.C.  - NADPH--hemoprotein reductase.
[IntEnz]   [ExPASy]   [KEGG]   [BRENDA]
      Reaction: NADPH + n oxidized hemoprotein = NADP+ + n reduced hemoprotein
Bound ligand (Het Group name = NAP)
corresponds exactly
+ n oxidized hemoprotein
= NADP(+)
+ n reduced hemoprotein
      Cofactor: FAD; FMN
Bound ligand (Het Group name = FAD) corresponds exactly
Molecule diagrams generated from .mol files obtained from the KEGG ftp site
 Gene Ontology (GO) functional annotation 
  GO annot!
  Cellular component     membrane   6 terms 
  Biological process     regulation of growth plate cartilage chondrocyte proliferation   24 terms 
  Biochemical function     electron carrier activity     13 terms  


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.
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.  

Literature references that cite this PDB file's key reference

  PubMed id Reference
20857167 D.Degregorio, S.J.Sadeghi, G.Di Nardo, G.Gilardi, and S.P.Solinas (2011).
Understanding uncoupling in the multiredox centre P450 3A4-BMR model system.
  J Biol Inorg Chem, 16, 109-116.  
20940534 D.Sandee, K.Morrissey, V.Agrawal, H.K.Tam, M.A.Kramer, T.S.Tracy, K.M.Giacomini, and W.L.Miller (2010).
Effects of genetic variants of human P450 oxidoreductase on catalysis by CYP2D6 in vitro.
  Pharmacogenet Genomics, 20, 677-686.  
20697309 V.Agrawal, J.H.Choi, K.M.Giacomini, and W.L.Miller (2010).
Substrate-specific modulation of CYP3A4 activity by genetic variants of cytochrome P450 oxidoreductase.
  Pharmacogenet Genomics, 20, 611-618.  
19737939 C.Xia, I.Misra, T.Iyanagi, and J.J.Kim (2009).
Regulation of interdomain interactions by calmodulin in inducible nitric-oxide synthase.
  J Biol Chem, 284, 30708-30717.  
19171935 D.Hamdane, C.Xia, S.C.Im, H.Zhang, J.J.Kim, and L.Waskell (2009).
Structure and function of an NADPH-cytochrome P450 oxidoreductase in an open conformation capable of reducing cytochrome P450.
  J Biol Chem, 284, 11374-11384.
PDB code: 3es9
19222033 E.Beaumont, J.C.Lambry, M.Blanchard-Desce, P.Martasek, S.P.Panda, E.E.van Faassen, J.C.Brochon, E.Deprez, and A.Slama-Schwok (2009).
NO formation by neuronal NO-synthase can be controlled by ultrafast electron injection from a nanotrigger.
  Chembiochem, 10, 690-701.  
19858215 J.Ellis, A.Gutierrez, I.L.Barsukov, W.C.Huang, J.G.Grossmann, and G.C.Roberts (2009).
Domain motion in cytochrome P450 reductase: conformational equilibria revealed by NMR and small-angle x-ray scattering.
  J Biol Chem, 284, 36628-36637.  
18487202 R.P.Ilagan, M.Tiso, D.W.Konas, C.Hemann, D.Durra, R.Hille, and D.J.Stuehr (2008).
Differences in a conformational equilibrium distinguish catalysis by the endothelial and neuronal nitric-oxide synthase flavoproteins.
  J Biol Chem, 283, 19603-19615.  
18433346 S.N.Hart, and X.B.Zhong (2008).
P450 oxidoreductase: genetic polymorphisms and implications for drug metabolism and toxicity.
  Expert Opin Drug Metab Toxicol, 4, 439-452.  
18551037 V.Agrawal, N.Huang, and W.L.Miller (2008).
Pharmacogenetics of P450 oxidoreductase: effect of sequence variants on activities of CYP1A2 and CYP2C19.
  Pharmacogenet Genomics, 18, 569-576.  
17534532 A.W.Munro, H.M.Girvan, and K.J.McLean (2007).
Variations on a (t)heme--novel mechanisms, redox partners and catalytic functions in the cytochrome P450 superfamily.
  Nat Prod Rep, 24, 585-609.  
17635179 C.E.Flück, C.Nicolo, and A.V.Pandey (2007).
Clinical, structural and functional implications of mutations and polymorphisms in human NADPH P450 oxidoreductase.
  Fundam Clin Pharmacol, 21, 399-410.  
18028029 K.J.McLean, H.M.Girvan, and A.W.Munro (2007).
Cytochrome P450/redox partner fusion enzymes: biotechnological and toxicological prospects.
  Expert Opin Drug Metab Toxicol, 3, 847-863.  
16642502 J.Hritz, G.Zoldák, and E.Sedlák (2006).
Cofactor assisted gating mechanism in the active site of NADH oxidase from Thermus thermophilus.
  Proteins, 64, 465-476.  
16249336 M.Jáchymová, P.Martásek, S.Panda, L.J.Roman, M.Panda, T.M.Shea, Y.Ishimura, J.J.Kim, and B.S.Masters (2005).
Recruitment of governing elements for electron transfer in the nitric oxide synthase family.
  Proc Natl Acad Sci U S A, 102, 15833-15838.  
16080215 P.Meinhold, M.W.Peters, M.M.Chen, K.Takahashi, and F.H.Arnold (2005).
Direct conversion of ethane to ethanol by engineered cytochrome P450 BM3.
  Chembiochem, 6, 1765-1768.  
15182370 A.J.Dunford, K.R.Marshall, A.W.Munro, and N.S.Scrutton (2004).
Thermodynamic and kinetic analysis of the isolated FAD domain of rat neuronal nitric oxide synthase altered in the region of the FAD shielding residue Phe1395.
  Eur J Biochem, 271, 2548-2560.  
12787027 A.Gutierrez, A.W.Munro, A.Grunau, C.R.Wolf, N.S.Scrutton, and G.C.Roberts (2003).
Interflavin electron transfer in human cytochrome P450 reductase is enhanced by coenzyme binding. Relaxation kinetic studies with coenzyme analogues.
  Eur J Biochem, 270, 2612-2621.  
12567183 D.Leys, J.Basran, F.Talfournier, M.J.Sutcliffe, and N.S.Scrutton (2003).
Extensive conformational sampling in a ternary electron transfer complex.
  Nat Struct Biol, 10, 219-225.
PDB codes: 1o94 1o95 1o96 1o97
14653815 G.Zoldák, R.Sut'ák, M.Antalík, M.Sprinzl, and E.Sedlák (2003).
Role of conformational flexibility for enzymatic activity in NADH oxidase from Thermus thermophilus.
  Eur J Biochem, 270, 4887-4897.  
12631275 R.D.Finn, J.Basran, O.Roitel, C.R.Wolf, A.W.Munro, M.J.Paine, and N.S.Scrutton (2003).
Determination of the redox potentials and electron transfer properties of the FAD- and FMN-binding domains of the human oxidoreductase NR1.
  Eur J Biochem, 270, 1164-1175.  
12359874 S.Adak, M.Sharma, A.L.Meade, and D.J.Stuehr (2002).
A conserved flavin-shielding residue regulates NO synthase electron transfer and nicotinamide coenzyme specificity.
  Proc Natl Acad Sci U S A, 99, 13516-13521.  
The most recent references are shown first. Citation data come partly from CiteXplore and partly from an automated harvesting procedure. Note that this is likely to be only a partial list as not all journals are covered by either method. However, we are continually building up the citation data so more and more references will be included with time. Where a reference describes a PDB structure, the PDB code is shown on the right.