Glutamate-haem ester bond formation is disfavoured in flavocytochrome P450 BM3: Characterization of glutamate substitution mutants at the haem site of P450 BM3

Biochemical Journal

Volumn 427, Issue 3, 2010, Pages 455-466

  Girvan, Hazel M ^a   Levy, Colin W ^a   Williams, Paul ^a   Fisher, Karl ^a   Cheesman, Myles R ^b   Rigby, Stephen E J ^a   Leys, David ^a   Munro, Andrew W ^a  

^a UNIVERSITY OF MANCHESTER   (United Kingdom)

^b UNIVERSITY OF EAST ANGLIA   (United Kingdom)

Author keywords

Bacillus megaterium; Cytochrome P450; Electron transfer; Ferrous oxy complex; Flavocytochrome; Substrate oxidation

Indexed keywords

BACILLUS MEGATERIUM; CYTOCHROME P450; ELECTRON TRANSFER; FLAVOCYTOCHROME; SUBSTRATE OXIDATION;

ACIDS; ANTIFERROMAGNETISM; BACTERIOLOGY; BIODEGRADATION; CARBOXYLATION; COMPLEXATION; ELECTRON SPIN RESONANCE SPECTROSCOPY; ELECTRON TRANSITIONS; ESTERIFICATION; ESTERS; FATTY ACIDS; HYDROCARBONS; IRON; IRON COMPOUNDS; OXIDATION; OXYGEN; STRUCTURAL ANALYSIS; SUBSTRATES;

FREE RADICAL REACTIONS;

ARACHIDONIC ACID; CARBENE; CARBOXYLIC ACID DERIVATIVE; CYTOCHROME P450 BM3; DECANOIC ACID DERIVATIVE; ESTER; FERRIC ION; FERROUS ION; GLUTAMIC ACID; HEME; LAURIC ACID; METHYL GROUP; OMEGA 1 HYDROXYDODECANOATE; OMEGA 3 HYDROXYDODECANOATE; SUPEROXIDE; UNCLASSIFIED DRUG; BACTERIAL PROTEIN; CYTOCHROME P450; FLAVOCYTOCHROME P450 BM3 MONOXYGENASES; LAURIC ACID DERIVATIVE; REDUCED NICOTINAMIDE ADENINE DINUCLEOTIDE PHOSPHATE FERRIHEMOPROTEIN REDUCTASE;

AMINO ACID SUBSTITUTION; ARTICLE; BIOENGINEERING; CATALYSIS; CONTROLLED STUDY; COVALENT BOND; CRYSTALLOGRAPHY; ELECTRON SPIN RESONANCE; ELECTRON TRANSPORT; HYDROGEN BOND; HYDROXYLATION; MUTANT; MUTATION; NONHUMAN; PLASTICITY; POTENTIOMETRY; PRIORITY JOURNAL; PROTEIN DOMAIN; RAMAN SPECTROMETRY; SPECTROSCOPY; STRUCTURE ANALYSIS; THERMODYNAMICS; ULTRAVIOLET SPECTROSCOPY; WILD TYPE; CHEMISTRY; ENZYME SPECIFICITY; GENETICS; KINETICS; METABOLISM; PHYSIOLOGY; PROTEIN TERTIARY STRUCTURE; SITE DIRECTED MUTAGENESIS; ULTRAVIOLET SPECTROPHOTOMETRY;

BACILLUS MEGATERIUM; EUKARYOTA;

ARACHIDONIC ACID; BACTERIAL PROTEINS; CRYSTALLOGRAPHY; CYTOCHROME P-450 ENZYME SYSTEM; ELECTRON SPIN RESONANCE SPECTROSCOPY; ESTERS; GLUTAMIC ACID; HEME; KINETICS; LAURIC ACIDS; MUTAGENESIS, SITE-DIRECTED; MUTATION; NADPH-FERRIHEMOPROTEIN REDUCTASE; POTENTIOMETRY; PROTEIN STRUCTURE, TERTIARY; SPECTROPHOTOMETRY, ULTRAVIOLET; SPECTRUM ANALYSIS, RAMAN; SUBSTRATE SPECIFICITY;

EID: 77951241302     PISSN: 02646021     EISSN: 14708728     Source Type: Journal    
DOI: 10.1042/BJ20091603     Document Type: Article

References (55)

1. Denisov, I. G., Makris, T. M., Sligar, S. G. and Schlichting, I. (2005) Structure and chemistry of cytochrome P450. Chem. Rev. 105, 2253-2277
2. Guengerich, F. P., Wu, Z. L. and Bartleson, C. J. (2005) Function of human cytochrome P450s: characterization of the orphans. Biochem. Biophys. Res. Commun. 338, 465-469
3. Cupp-Vickery, J. R., Han, O., Hutchinson, C. R. and Poulos, T. L. (1996) Substrate-assisted catalysis in cytochrome P450eryF. Nat. Struct. Biol. 3, 632-637
4. Lawson, R. J., Leys, D., Sutcliffe, M. J., Kemp, C. A., Cheesman, M. R., Smith, S. J., Clarkson, J., Smith, W. E., Haq, I., Perkins, J. B. and Munro, A. W. (2004) Thermodynamic and biophysical characerterization of cytochrome P450 Biol from Bacillus subtilis. Biochemistry 43, 12410-12426
5. Poulos, T. L. (2003) The past and present of P450cam structural biology. Biochem. Biophys. Res. Commun. 312, 35-39
6. Munro, A. W., Leys, D. G., McLean, K. J., Marshall, K. R., Ost, T. W., Daff, S., Miles, C. S., Chapman, S. K., Lysek, D. A., Moser, C. C. et al. (2002) P450 BM3: the very model of a modern flavocytochrome. Trends Biochem. Sci. 27, 250-257
7. Perera, R., Sono, M., Sigman, J. A., Pfister, T. D., Lu, Y. and Dawson, J. H. (2003) Neutral thiol as a proximal ligand to ferrous heme iron: implications for heme proteins that lose cysteine thiolate ligation on reduction. Proc. Natl. Acad. Sci. U.S.A. 100, 3641-3646
8. Munro, A. W., Girvan, H. M. and McLean, K. J. (2007) Cytochrome P450-redox partner fusion enzymes. Biochim. Biophys. Acta 1770, 345-359
9. Munro, A. W., Girvan, H. M. and McLean, K. J. (2007) Variations on a (t)heme: novel mechanisms, redox partners and catalytic functions in the cytochrome P450 superfamily. Nat. Prod. Rep. 24, 585-609
10. Colas, C. and Ortiz de Montellano, P. R. (2003) Autocatalytic radical reactions in physiological prosthetic heme modification. Chem. Rev. 103, 2305-2332
11. Henne, K. R., Kunze, K. L., Zheng, Y. M., Christmas, P., Soberman, R. J. and Rettie, A. E. (2001) Covalent linkage of prosthetic heme to CYP4 family enzymes. Biochemistry 40, 12925-12931
12. LeBrun, L. A., Hoch, U. and Ortiz de Montellano, P. R. (2002) Autocatalytic mechanism and consequences of covalent heme attachment in the cytochrome P4504A family. J. Biol. Chem. 277, 12755-12761
13. Hoch, U. and Ortiz de Montellano, P. R. (2001) Covalently linked heme in cytochrome P4504A fatty acid hydroxylases. J. Biol. Chem. 276, 11339-11346
14. Limburg, J., LeBrun, L. A. and Ortiz de Montellano, P. R. (2005) The P450cam G248E mutant covalently binds its prosthetic heme group. Biochemistry 44, 4091-4099
15. Girvan, H. M., Marshall, K. R., Lawson, R. J., Leys, D., Joyce, M. G., Clarkson, J., Smith, W. E., Cheesman, M. R. and Munro, A. W. (2004) Flavocytochrome P450 BM3 mutant A264E undergoes substrate-dependent formation of a novel heme iron ligand set. J. Biol. Chem. 279, 23274-23286
16. Girvan, H. M., Seward, H. E., Toogood, H. S., Cheesman, M. R., Leys, D. and Munro, A. W. (2007) Structural and spectroscopic characterization of P450 BM3 mutants with unprecedented P450 heme iron ligand sets: new heme ligation states influence conformational equilibria in P450 BM3. J. Biol. Chem. 282, 564-572
17. Girvan, H. M., Toogood, H. S., Littleford, R. E., Seward, H. E., Smith, W. E., Ekanem, I. S., Leys, D., Cheesman, M. R. and Munro, A. W. (2009) Novel haem co-ordination variants of flavocytochrome P450 BM3. Biochem. J. 417, 65-76
18. 268 in oxygen activation of cytochrome P450 BM3. Biochemistry 34, 14733-14740
19. Ost, T. W., Miles, C. S., Munro, A. W., Murdoch, J., Reid, G. A. and Chapman, S. K. (2001) Phenylalanine 393 exerts thermodynamic control over the heme of flavocytochrome P450 BM3. Biochemistry 40, 13421-13429
20. Ravichandran, K. G., Boddupalli, S. S., Hasemann, C. A., Peterson, J. A. and Deisenhofer, J. (1993) Crystal structure of hemoprotein domain of P450BM-3, a prototype for microsomal P450s. Science 261, 731-736
21. Whitehouse, J. C., Bell, S. G., Yang, W., Yorke, J. A., Blanford, C. F., Strong, A. J. F., Morse, E. J., Bartlam, M., Rao, Z. and Wong, L.-L. (2009) A highly active single-mutation variant of P450 BM3 (CYP102A1). ChemBioChem 10, 1654-1656
22. Miles, J. S., Munro, A. W., Rospendowski, B. N., Smith, W. E., McKnight, J. and Thomson, A. J. (1992) Domains of the catalytically self-sufficient cytochrome P-450 BM-3: genetic construction, overexpression, purification and spectroscopic characterization. Biochem. J. 288, 503-509
23. Noble, M. A., Miles, C. S., Chapman, S. K., Lysek, D. A., MacKay, A. C., Reid, G. A., Hanzlik, R. P. and Munro, A. W. (1999) Roles of key active-site residues in flavocytochrome P450 BM3. Biochem. J. 339, 371-379
24. Omura, T. and Sato, R. (1964) The carbon monoxide-binding pigment of liver microsome. I. Evidence for its hemoprotein nature. J. Biol. Chem. 239, 2370-2378
25. McLean, K. J., Cheesman, M. R., Rivers, S. L., Richmond, A., Leys, D., Chapman, S. K., Reid, G. A., Price, N. C., Kelly, S. M., Clarkson, J. et al. (2002) Expression, purification and spectroscopic characterization of the cytochrome P450 CYP121 from Mycobacterium tuberculosis. J. Inorg. Biochem. 91, 527-541
26. Dutton, P. L. (1978) Redox potentiometry: determination of midpoint potentials of oxidation-reduction components of biological electron-transfer systems. Methods Enzymol. 54, 411-435
27. Daff, S. N., Chapman, S. K., Turner, K. L., Holt, R. A., Govindaraj, S., Poulos, T. L. and Munro, A. W. (1997) Redox control of the catalytic cycle of flavocytochrome P-450 BM3. Biochemistry 36, 13816-13823
28. Kabsch, W. (1988) Automatic indexing of rotation diffraction patterns. J. Appl. Crystallogr. 21, 67-71
29. Collaborative Computational Project, Number 4 (1994) The CCP4 suite: programs for protein crystallography. Acta Crystallogr. Sect. D Biol. Crystallogr. 50, 760-763
30. Murshudov, G. N., Vagin, A. A. and Dodson, E. J. (1997) Refinement of macromolecular structures by the maximum-likelihood method. Acta Crystallogr. Sect. D Biol. Crystallogr. 53, 240-255
31. Emsley, P. and Cowtan, K. (2004) Coot: model-building tools for molecular graphics. Acta Crystallogr. Sect. D Biol. Crystallogr. 60, 2126-2132
32. Joyce, M. G., Girvan, H. M., Munro, A. W. and Leys, D. (2004) A single mutation in cytochrome P450 BM3 induces the conformational rearrangement seen upon substrate binding in the wild-type enzyme. J. Biol. Chem. 279, 23287-23293
33. Li, H. Y. and Poulos, T. L. (1997) The structure of the cytochrome P450BM-3 haem domain complexed with the fatty acid substrate, palmitoleic acid. Nat. Struct. Biol. 4, 140-146
34. Perera, R., Sono, M., Sigman, J. A., Pfister, T. D., Lu, Y. and Dawson, J. H. (2003) Neutral thiol as a proximal ligand to ferrous heme iron: implications for heme proteins that lose cysteine thiolate ligation on reduction. Proc. Natl. Acad. Sci. U.S.A. 100, 3641-3646
35. Dunford, A. J., McLean, K. J., Sabri, M., Seward, H. E., Heyes, D. J., Scrutton, N. S. and Munro, A. W. (2007) Rapid P450 heme iron reduction by laser photoexcitation of Mycobacterium tuberculosisCYP121 and CYP51B1: analysis of CO complexation reactions and reversibility of the P450/P420 equilibrium. J. Biol. Chem. 282, 24816-24824
36. Hu, S., Smith, K. M. and Spiro, T. G. (1996) Assignment of protoheme resonance Raman spectrum by heme labelling in myoglobin. J. Am. Chem. Soc. 118, 12638-12646
37. Smith, S. J., Munro, A. W. and Smith, W. E. (2003) Resonance Raman scattering of cytochrome P450 BM3 and the effect of imidazole inhibitors. Biopolymers 70, 620-627
38. Ost, T. W. B., Munro, A. W., Mowat, C. G., Taylor, P. R., Pesseguiero, A., Fulco, A. J., Cho, A. K., Cheesman, M. R., Walkinshaw, M. D. and Chapman, S. K. (2001) Structural and spectroscopic analysis of the F393H mutant of flavocytochrome P450 BM3. Biochemistry 40, 13430-13438 (Pubitemid 33130770)
39. Munro, A. W., Daff, S., Coggins, J. R., Lindsay, J. G. and Chapman, S. K. (1996) Probing electron transfer in flavocytochrome P450 BM3 and its component domains. Eur. J. Biochem. 239, 403-409
40. Girvan, H. M., Heyes, D. J., Scrutton, N. S. and Munro, A. W. (2007) Laser photoexcitation of NAD(P)H induces reduction of P450 BM3 heme domain on the microsecond time scale. J. Am. Chem. Soc. 129, 6647-6653
41. Hanley, S. C. and Daff, S. (2004) The unusual redox properties of flavocytochrome P450 BM3 flavodoxin domain. Biochem. Biophys. Res. Commun. 325, 1418-1423
42. Sevrioukova, I. F. and Peterson, J. A. (1995) Reaction of carbon monoxide and molecular oxygen with P450terp (CYP108) and P450BM-3 (CYP102). Arch. Biochem. Biophys. 317, 397-404
43. Bec, N., Anzenbacher, P., Anzenbacherova, E., Gorren, A. C. F., Munro, A. W. and Lange, R. (1999) Spectral properties of the oxyferrous complex of the heme domain of cytochrome P450 BM-3 (CYP102). Biochem. Biophys. Res. Commun. 266, 187-189
44. Davydov, R., Makris, T. M., Kofman, V., Werst, D. E., Sligar, S. G. and Hoffman, B. M. (2001) Hydroxylation of camphor by reduced oxy-cytochrome P450cam: mechanistic implications of EPR and ENDOR studies of catalytic intermediates in native and mutant enzyme. J. Am. Chem. Soc. 123, 1403-1415
45. Gantt, S., Denisov, I. G., Grinkova, Y. V. and Sligar, S. G. (2009) The critical iron-oxygen intermediate in human aromatase. Biochem. Biophys. Res. Commun. 387, 169-173
46. Boddupalli, S. S., Pramanik, B. C., Slaughter, C. A., Estabrook, R. W. and Peterson, J. A. (1992) Fatty acid monooxygenation by P450BM-3: product identification and proposed mechanisms for the sequential hydroxylation reactions. Arch. Biochem. Biophys. 292, 20-28
47. Haines, D. C., Tomchick, D. R., Machius, M. and Peterson, J. A. (2001) Pivotal role of water in the mechanism of P450BM-3. Biochemistry 40, 13456-13465
48. Sevrioukova, I. F., Li, H., Zhang, H., Peterson, J. A. and Poulos, T. L. (1999) Structure of a cytochrome P450-redox partner electron-transfer complex. Proc. Natl. Acad. Sci. U.S.A. 96, 1863-1868
49. Ost, T. W., Miles, C. S., Murdoch, J., Cheung, Y., Reid, G. A., Chapman, S. K. and Munro, A. W. (2000) Rational re-design of the substrate binding site of flavocytochrome P450 BM3. FEBS Lett. 486, 173-177
50. Neeli, R., Girvan, H. M., Lawrence, A., Warren, M. J., Leys, D., Scrutton, N. S. and Munro, A. W. (2005) The dimeric form of flavocytochrome P450 BM3 is catalytically functional as a fatty acid hydroxylase. FEBS Lett. 579, 5582-5588
51. Lawson, R. J., Leys, D., Sutcliffe, M. J., Kemp, C. A., Cheesman, M. R., Smith, S. J., Smith, W. E., Haq, I., Perkins, J. B. and Munro, A. W. (2004) Thermodynamic and biophysical characterization of cytochrome P450 Biol from Bacillus subtilis. Biochemistry 43, 12410-12426
52. Ost, T. W., Clark, J., Mowat, C. G., Miles, C. S., Walkinshaw, M. D., Reid, G. A., Chapman, S. K. and Daff, S. (2003) Oxygen activation and electron transfer in flavocytochrome P450 BM3. J. Am. Chem. Soc. 125, 15010-15020
53. Xu, F., Bell, S. G., Lednik, J., Insley, A., Rao, Z. and Wong, L.-L. (2005) The heme monooxygenase cytochrome P450cam can be engineered to oxidize ethane to ethanol. Angew. Chem. Int. Ed. Engl. 44, 4029-4032
54. Sharrock, M., Debrunner, P. G., Schulz, C., Lipscomb, J. D., Marshall, V. and Gunsalus, I. C. (1976) Cytochrome P450cam and its complexes: Mossbauer parameters of the heme iron. Biochim. Biophys. Acta 420, 8-26
55. Bangcharoenpaurpong, O., Rizos, A. K., Champion, P. M., Jollie, D. and Sligar, S. G. (1986) Resonance Raman detection of bound dioxygen in cytochrome P-450cam. J. Biol. Chem. 261, 8089-8092