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PDBsum entry 1yqp

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protein ligands Protein-protein interface(s) links
Oxidoreductase PDB id
1yqp
Jmol
Contents
Protein chains
441 a.a. *
Ligands
HEM ×2
Waters ×973
* Residue conservation analysis
PDB id:
1yqp
Name: Oxidoreductase
Title: T268n mutant cytochrome domain of flavocytochrome p450 bm3
Structure: Bifunctional p-450:nadph-p450 reductase. Chain: a, b. Fragment: cytochrome domain. Synonym: cytochrome p450(bm-3), p450bm-3. Engineered: yes. Mutation: yes
Source: Bacillus megaterium. Organism_taxid: 1404. Expressed in: escherichia coli. Expression_system_taxid: 562.
Resolution:
1.80Å     R-factor:   0.176     R-free:   0.210
Authors: J.P.Clark,C.S.Miles,C.G.Mowat,M.D.Walkinshaw,G.A.Reid, S.N.Daff,S.K.Chapman
Key ref: J.P.Clark et al. (2006). The role of Thr268 and Phe393 in cytochrome P450 BM3. J Inorg Biochem, 100, 1075-1090. PubMed id: 16403573 DOI: 10.1016/j.jinorgbio.2005.11.020
Date:
02-Feb-05     Release date:   31-Jan-06    
PROCHECK
Go to PROCHECK summary
 Headers
 References

Protein chains
Pfam   ArchSchema ?
P14779  (CPXB_BACME) -  Bifunctional P-450/NADPH-P450 reductase
Seq:
Struc:
 
Seq:
Struc:
 
Seq:
Struc:
1049 a.a.
441 a.a.*
Key:    PfamA domain  Secondary structure  CATH domain
* PDB and UniProt seqs differ at 1 residue position (black cross)

 Enzyme reactions 
   Enzyme class 2: E.C.1.14.14.1  - Unspecific monooxygenase.
[IntEnz]   [ExPASy]   [KEGG]   [BRENDA]
      Reaction: RH + reduced flavoprotein + O2 = ROH + oxidized flavoprotein + H2O
RH
+ reduced flavoprotein
+ O(2)
= ROH
+ oxidized flavoprotein
+ H(2)O
      Cofactor: Heme-thiolate
   Enzyme class 3: E.C.1.6.2.4  - NADPH--hemoprotein reductase.
[IntEnz]   [ExPASy]   [KEGG]   [BRENDA]
      Reaction: NADPH + n oxidized hemoprotein = NADP+ + n reduced hemoprotein
NADPH
+ n oxidized hemoprotein
= NADP(+)
+ n reduced hemoprotein
      Cofactor: FAD; FMN
FAD
FMN
Note, where more than one E.C. class is given (as above), each may correspond to a different protein domain or, in the case of polyprotein precursors, to a different mature protein.
Molecule diagrams generated from .mol files obtained from the KEGG ftp site
 Gene Ontology (GO) functional annotation 
  GO annot!
  Biological process     oxidation-reduction process   1 term 
  Biochemical function     oxidoreductase activity, acting on paired donors, with incorporation or reduction of molecular oxygen     3 terms  

 

 
    reference    
 
 
DOI no: 10.1016/j.jinorgbio.2005.11.020 J Inorg Biochem 100:1075-1090 (2006)
PubMed id: 16403573  
 
 
The role of Thr268 and Phe393 in cytochrome P450 BM3.
J.P.Clark, C.S.Miles, C.G.Mowat, M.D.Walkinshaw, G.A.Reid, S.N.Daff, S.K.Chapman.
 
  ABSTRACT  
 
In flavocytochrome P450 BM3 there are several active site residues that are highly conserved throughout the P450 superfamily. Of these, a phenylalanine (Phe393) has been shown to modulate heme reduction potential through interactions with the implicitly conserved heme-ligand cysteine. In addition, a distal threonine (Thr268) has been implicated in a variety of roles including proton donation, oxygen activation and substrate recognition. Substrate binding in P450 BM3 causes a shift in the spin state from low- to high-spin. This change in spin-state is accompanied by a positive shift in the reduction potential (DeltaE(m) [WT+arachidonate (120 microM)]=+138 mV). Substitution of Thr268 by an alanine or asparagine residue causes a significant decrease in the ability of the enzyme to generate the high-spin complex via substrate binding and consequently leads to a decrease in the substrate-induced potential shift (DeltaE(m) [T268A+arachidonate (120 microM)]=+73 mV, DeltaE(m) [T268N+arachidonate (120 microM)]=+9 mV). Rate constants for the first electron transfer and for oxy-ferrous decay were measured by pre-steady-state stopped-flow kinetics and found to be almost entirely dependant on the heme reduction potential. More positive reduction potentials lead to enhanced rate constants for heme reduction and more stable oxy-ferrous species. In addition, substitutions of the threonine lead to an increase in the production of hydrogen peroxide in preference to hydroxylated product. These results suggest an important role for this active site threonine in substrate recognition and in maintaining an efficiently functioning enzyme. However, the dependence of the rate constants for oxy-ferrous decay on reduction potential raises some questions as to the importance of Thr268 in iron-oxo stabilisation.
 

Literature references that cite this PDB file's key reference

  PubMed id Reference
21154803 M.Ma, S.G.Bell, W.Yang, Y.Hao, N.H.Rees, M.Bartlam, W.Zhou, L.L.Wong, and Z.Rao (2011).
Structural Analysis of CYP101C1 from Novosphingobium aromaticivorans DSM12444.
  Chembiochem, 12, 88-99.
PDB codes: 3oft 3ofu
21110374 C.J.Whitehouse, W.Yang, J.A.Yorke, B.C.Rowlatt, A.J.Strong, C.F.Blanford, S.G.Bell, M.Bartlam, L.L.Wong, and Z.Rao (2010).
Structural basis for the properties of two single-site proline mutants of CYP102A1 (P450BM3).
  Chembiochem, 11, 2549-2556.
PDB code: 3m4v
20446763 T.C.Pochapsky, S.Kazanis, and M.Dang (2010).
Conformational plasticity and structure/function relationships in cytochromes P450.
  Antioxid Redox Signal, 13, 1273-1296.  
17957765 E.Stjernschantz, B.M.van Vugt-Lussenburg, A.Bonifacio, S.B.de Beer, G.van der Zwan, C.Gooijer, J.N.Commandeur, N.P.Vermeulen, and C.Oostenbrink (2008).
Structural rationalization of novel drug metabolizing mutants of cytochrome P450 BM3.
  Proteins, 71, 336-352.  
18815130 J.Tejero, A.Biswas, Z.Q.Wang, R.C.Page, M.M.Haque, C.Hemann, J.L.Zweier, S.Misra, and D.J.Stuehr (2008).
Stabilization and Characterization of a Heme-Oxy Reaction Intermediate in Inducible Nitric-oxide Synthase.
  J Biol Chem, 283, 33498-33507.
PDB code: 3dwj
18431595 K.M.Pajerowska-Mukhtar, M.S.Mukhtar, N.Guex, V.A.Halim, S.Rosahl, I.E.Somssich, and C.Gebhardt (2008).
Natural variation of potato allene oxide synthase 2 causes differential levels of jasmonates and pathogen resistance in Arabidopsis.
  Planta, 228, 293-306.  
18161730 M.J.Cryle, and J.J.De Voss (2008).
The role of the conserved threonine in P450 BM3 oxygen activation: substrate-determined hydroxylation activity of the Thr268Ala mutant.
  Chembiochem, 9, 261-266.  
18473391 R.J.Branco, A.Seifert, M.Budde, V.B.Urlacher, M.J.Ramos, and J.Pleiss (2008).
Anchoring effects in a wide binding pocket: the molecular basis of regioselectivity in engineered cytochrome P450 monooxygenase from B. megaterium.
  Proteins, 73, 597-607.  
17318599 A.Bonifacio, A.R.Groenhof, P.H.Keizers, C.de Graaf, J.N.Commandeur, N.P.Vermeulen, A.W.Ehlers, K.Lammertsma, C.Gooijer, and G.van der Zwan (2007).
Altered spin state equilibrium in the T309V mutant of cytochrome P450 2D6: a spectroscopic and computational study.
  J Biol Inorg Chem, 12, 645-654.  
17650504 I.G.Denisov, Y.V.Grinkova, M.A.McLean, and S.G.Sligar (2007).
The one-electron autoxidation of human cytochrome P450 3A4.
  J Biol Chem, 282, 26865-26873.  
16762915 I.G.Denisov, Y.V.Grinkova, B.J.Baas, and S.G.Sligar (2006).
The ferrous-dioxygen intermediate in human cytochrome P450 3A4. Substrate dependence of formation and decay kinetics.
  J Biol Chem, 281, 23313-23318.  
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 codes are shown on the right.