P-LinK: A method for generating multicomponent cytochrome P450 fusions with variable linker length

BioTechniques

Volumn 57, Issue 1, 2014, Pages 13-20

  Belsare, Ketaki D ^a   Joëlle Ruff, Anna ^a   Martinez, Ronny ^a   Shivange, Amol V ^b   Mundhada, Hemanshu ^c   Holtmann, Dirk ^d   Schrader, Jens ^d   Schwaneberg, Ulrich ^a  

^a RWTH AACHEN UNIVERSITY   (Germany)

^b University of California   (United States)

^c Chalmers University of Technology   (Denmark)

^d KARL WINNACKER INSTITUT   (Germany)

Author keywords

Directed evolution; Fusion protein; Linker; Monooxygenase; Multicomponent system; PLICing; Protein engineering

Indexed keywords

CYTOCHROME P450; FLAVINE ADENINE NUCLEOTIDE; FLAVINE MONONUCLEOTIDE; FLAVODOXIN; HYBRID PROTEIN; UNSPECIFIC MONOOXYGENASE;

ARTICLE; BACTERIAL GENOME; ENZYME ACTIVITY; ESCHERICHIA COLI; GAS CHROMATOGRAPHY; HUMAN; MOLECULAR CLONING; NUCLEOTIDE SEQUENCE; POLYMERASE CHAIN REACTION; PROTEIN CONSTRUCTION; PROTEIN ENGINEERING; PROTEIN EXPRESSION; PROTEIN PURIFICATION; SEQUENCE ANALYSIS; THIN LAYER CHROMATOGRAPHY; AMINO ACID SEQUENCE; CHEMISTRY; GENE LIBRARY; GENETICS; METABOLISM; MOLECULAR GENETICS; PROCEDURES;

AMINO ACID SEQUENCE; CYTOCHROME P-450 ENZYME SYSTEM; ESCHERICHIA COLI; GENE LIBRARY; MOLECULAR SEQUENCE DATA; POLYMERASE CHAIN REACTION; PROTEIN ENGINEERING; RECOMBINANT FUSION PROTEINS;

EID: 84904094364     PISSN: 07366205     EISSN: 19409818     Source Type: Journal    
DOI: 10.2144/000114187     Document Type: Article

References (37)

1. Clark, E.D.B. 2001. Protein refolding for industrial processes. Curr. Opin. Biotechnol. 12: 202-207.
2. Georgiou, G., C. Stathopoulos, P.S. Daugherty, A.R. Nayak, B.L. Iverson, and R. Curtiss III. 1997. Display of heterologous proteins on the surface of microorganisms: from the screening of combinatorial libraries to live recombinant vaccines. Nat. Biotechnol. 15: 29-34.
3. Schmidt, S.R. 2009. Fusion-proteins as biopharmaceuticals--applications and challenges. Curr Opin Drug Discov Devel. 12: 284-295.
4. Tychopoulos, M., L. Corcos, P. Genne, P. Beaune, and I. de Waziers. 2005. A virus-directed enzyme prodrug therapy (VDEPT) strategy for lung cancer using a CYP2B6/NADPH-cytochrome P450 reductase fusion protein. Cancer Gene Ther. 12: 497-508.
5. Uhlén, M., G. Forsberg, T. Moks, M. Hartmanis, and B. Nilsson. 1992. Fusion proteins in Biotechnology. Curr. Opin. Biotechnol. 3: 363-369.
6. Fasan, R. 2012. Tuning P450 Enzymes as Oxidation Catalysts. ACS Catal. 2: 647-666.
7. Gilardi, G., Y.T. Meharenna, G.E. Tsotsou, S.J. Sadeghi, M. Fairhead, and S. Giannini. 2002. Molecular Lego: design of molecular assemblies of P450 enzymes for nanobiotechnology. Biosens. Bioelectron. 17: 133-145.
8. Munro, A.W., H.M. Girvan, and K.J. McLean. 2007. Cytochrome P450-redox partner fusion enzymes. Biochim. Biophys. Acta 1770: 345-359.
9. Urlacher, V.B. and M. Girhard. 2012. Cytochrome P450 monooxygenases: an update on perspectives for synthetic application. Trends Biotechnol. 30: 26-36.
10. Whitehouse, C.J., S.G. Bell, and L.L. Wong. 2012. P450(BM3) (CYP102A1): Connecting the dots. Chem. Soc. Rev. 41: 1218-1260.
11. Sadeghi, S.J. and G. Gilardi. 2013. Chimeric P450 enzymes: Activity of artificial redox fusions driven by different reductases for biotechnological applications. Biotechnol. Appl. Biochem. 60: 102-110.
12. Choi, K.-Y., E. Jung, D.-H. Jung, B.-R. An, B.P. Pandey, H. Yun, C. Sung, H.-Y. Park, and B.-G. Kim. 2012. Engineering of daidzein 3'-hydroxylase P450 enzyme into catalytically self-sufficient cytochrome P450. Microb. Cell Fact. 11: 1-10.
13. Fisher, C.W., M.S. Shet, D.L. Caudle, C.A. Martin-Wixtrom, and R.W. Estabrook. 1992. High-level expression in Escherichia coli of enzymatically active fusion proteins containing the domains of mammalian cytochromes P450 and NADPH-P450 reductase flavoprotein. Proc. Natl. Acad. Sci. USA 89: 10817-10821.
14. Hirakawa, H. and T. Nagamune. 2010. Molecular assembly of P450 with ferredoxin and ferredoxin reductase by fusion to PCNA. ChemBioChem 11: 1517-1520.
15. Li, S., L.M. Podust, and D.H. Sherman. 2007. Engineering and analysis of a self-sufficient biosynthetic cytochrome P450 PikC fused to the RhFRED reductase domain. J. Am. Chem. Soc. 129: 12940-12941.
16. Robin, A., G.A. Roberts, J. Kisch, F. Sabbadin, G. Grogan, N. Bruce, N.J. Turner, and S.L. Flitsch. 2009. Engineering and improvement of the efficiency of a chimeric [P450cam-RhFRed reductase domain] enzyme. Chem Commun (Camb). 18: 2478-2480.
17. Shet, M.S., C.W. Fisher, P.L. Holmans, and R.W. Estabrook. 1993. Human cytochrome P450 3A4: enzymatic properties of a purified recombinant fusion protein containing NADPH-P450 reductase. Proc. Natl. Acad. Sci. USA 90: 11748-11752.
18. Sibbesen, O., J.J. De Voss, and P.R.O. de Montellano. 1996. Putidaredoxin reductase-putidaredoxin-cytochrome P450cam triple fusion protein construction of a self-sufficient Escherichia coli catalytic system. J. Biol. Chem. 271: 22462-22469.
19. Hirakawa, H., N. Kamiya, T. Tanaka, and T. Nagamune. 2007. Intramolecular electron transfer in a cytochrome P450cam system with a site-specific branched structure. Protein Eng. Des. Sel. 20: 453-459.
20. Nodate, M., M. Kubota, and N. Misawa. 2006. Functional expression system for cytochrome P450 genes using the reductase domain of self-sufficient P450RhF from Rhodococcus sp. NCIMB 9784. Appl. Microbiol. Biotechnol. 71: 455-462.
21. Govindaraj, S. and T.L. Poulos. 1995. Role of the linker region connecting the reductase and heme domains in cytochrome P450BM-3. Biochemistry 34: 11221-11226.
22. Govindaraj, S. and T.L. Poulos. 1996. Probing the structure of the linker connecting the reductase and heme domains of cytochrome P450BM-3 using site-directed mutagenesis. Protein Sci. 5: 1389-1393.
23. Blanusa, M., A. Schenk, H. Sadeghi, J. Marienhagen, and U. Schwaneberg. 2010. Phosphorothioate-based ligase-independent gene cloning (PLICing): An enzyme-free and sequence-independent cloning method. Anal. Biochem. 406: 141-146.
24. Zengin Çekiç, S., D. Holtmann, K. Mangold, U. Schwaneberg, and J. Schrader. 2010. Mediated Electron Transfer with P450cin. Electrochem. Commun. 12: 1547-1550.
25. Inoue, H., H. Nojima, and H. Okayama. 1990. High efficiency transformation of Escherichia coli with plasmids. Gene 96: 23-28.
26. Hawkes, D.B., G.W. Adams, A.L. Burlingame, P.R.O. de Montellano, and J.J. De Voss. 2002. Cytochrome P450cin (CYP176A), isolation, expression, and characterization. J. Biol. Chem. 277: 27725-27732.
27. Hawkes, D.B., K.E. Slessor, P.V. Bernhardt, and J.J. De Voss. 2010. Cloning, Expression and Purification of Cindoxin, an Unusual Fmn-Containing Cytochrome P450 Redox Partner. ChemBioChem 11: 1107-1114.
28. Jenkins, C.M. and M.R. Waterman. 1998. NADPH-flavodoxin reductase and flavodoxin from Escherichia coli: characteristics as a soluble microsomal P450 reductase. Biochemistry 37: 6106-6113.
29. Miyazawa, M. and Y. Hashimoto. 2002. Antimicrobial and bactericidal activities of esters of 2-endo-hydroxy-1, 8-cineole as new aroma chemicals. J. Agric. Food Chem. 50: 3522-3526.
30. Miyazawa, M., M. Shindo, and T. Shimada. 2001. Oxidation of 1, 8-cineole, the monoterpene cyclic ether originated from Eucalyptus polybractea, by cytochrome P450 3A enzymes in rat and human liver microsomes. Drug Metab. Dispos. 29: 200-205.
31. Meharenna, Y.T., H. Li, D.B. Hawkes, A.G. Pearson, J. De Voss, and T.L. Poulos. 2004. Crystal structure of P450cin in a complex with its substrate, 1, 8-cineole, a close structural homologue to D-camphor, the substrate for P450cam. Biochemistry 43: 9487-9494.
32. Chou, P.Y. and G.D. Fasman. 1974. Conformational parameters for amino acids in helical, b-sheet, and random coil regions calculated from proteins. Biochemistry 13: 211-222.
33. Chou, P.Y. and G.D. Fasman. 1974. Prediction of protein conformation. Biochemistry 13: 222-245.
34. George, R.A. and J. Heringa. 2002. An analysis of protein domain linkers: their classification and role in protein folding. Protein Eng. 15: 871-879.
35. Wriggers, W., S. Chakravarty, and P.A. Jennings. 2005. Control of protein functional dynamics by peptide linkers. Biopolymers. 80: 736-746.
36. Sevrioukova, I.F., H. Li, H. Zhang, J.A. Peterson, and T.L. Poulos. 1999. Structure of a cytochrome P450-redox partner electron-transfer complex. Proc. Natl. Acad. Sci. USA 96: 1863-1868.
37. Dennig, A., A.V. Shivange, J. Marienhagen, and U. Schwaneberg. 2011. OmniChange: the sequence independent method for simultaneous site-saturation of five codons. PLoS ONE 6: e26222.