Directed Evolution of Enzymes

Enzyme Catalysis in Organic Synthesis, Third Edition

Volumn 1, Issue , 2012, Pages 119-190

  Reetz, Manfred T ^a  

^a MAX PLANCK INSTITUT FÜR KOHLENFORSCHUNG   (Germany)

Author keywords

Directed evolution; Enzyme stability; Gene mutagenesis methods; Iterative saturation mutagenesis; Recombinant methods; Stereoselectivity

Indexed keywords

EID: 84872199976     PISSN: None     EISSN: None     Source Type: Book    
DOI: 10.1002/9783527639861.ch5     Document Type: Chapter

References (291)

1. Tao, J., Lin, G.-Q., and Liese, A. (2009) Biocatalysis for the Pharmaceutical Industry: Discovery, Development, and Manufacturing, John Wiley & Sons, Ltd., Chichester;
2. Gotor, V., Alfonso, I. and García-Urdiales, E. (eds) (2008) Asymmetric Organic Synthesis with Enzymes, Wiley-VCH Verlag GmbH, Weinheim;
3. Faber, K. (2004) Biotransformations in Organic Chemistry, 5th edn, Springer, Berlin;
4. Bommarius, A.S. and Riebel-Bommarius, B.R. (2004) Biocatalysis: Fundamentals and Applications, Wiley-VCH Verlag GmbH, Weinheim.
5. Lutz, S. and Bornscheuer, U.T. (2009) Protein Engineering Handbook, vols 1-2, Wiley-VCH Verlag GmbH, Weinheim.
6. Reetz, M.T. (2011) Laboratory evolution of stereoselective enzymes: A prolific source of catalysts for asymmetric reactions. Angew. Chem. Int. Ed., 50, 138-174;
7. Romero, P.A. and Arnold, F.H. (2009) Exploring protein fitness landscapes by directed evolution. Nat. Rev. Mol. Cell Biol., 10, 866-876;
8. Turner, N.J. (2009) Directed evolution drives the next generation of biocatalysts.Nat. Chem. Biol., 5, 567-573;
9. Jäckel, C., Kast, P. andHilvert, D. (2008) Protein design by directed evolution. Annu. Rev. Biophys. Biomol. Struct., 37, 153-173;
10. Bershtein, S. and Tawfik, D.S. (2008) Advances in laboratory evolution of enzymes. Curr. Opin. Chem. Biol., 12, 151-158;
11. Reetz, M.T. (2008) Directed evolution as a means to engineer enantioselective enzymes, in Asymmetric Organic Synthesis with Enzymes (eds V. Gotor, I. Alfonso, and E.García-Urdiales), Wiley-VCH Verlag GmbH, Weinheim, pp. 21-63;
12. Reetz, M.T. (2010) Enzyme engineering by directed evolution, in Manual of Industrial Microbiology and Biotechnology, 3rd edn (eds H. Zhao et al..) ASM Press, Washington D.C., pp. 466-479.
13. Mills, D.R., Peterson, R.L., and Spiegelman, S. (1967) An extracellular Darwinian experiment with a self-duplicating nucleic acid molecule. Proc. Natl. Acad. Sci. U.S.A., 58, 217-224.
14. Joyce, G.F. (2007) Forty years of in vitro evolution. Angew. Chem., 119, 6540-6557; Angew. Chem. Int. Ed., 46, 6420-6436.
15. Francis, J.C. and Hansche, P.E. (1972) Directed evolution of metabolic pathways in microbial populations. I. Modification of the acid phosphatase ph optimum in S. Cerevisiae. Genetics, 70, 59-73.
16. Hall, B.G. (1977) Number of mutations required to evolve a new lactase function in Escherichia coli. J. Bacteriol., 129, 540-543.
17. Eigen, M. and Gardiner, W. (1984) Evolutionary molecular engineering based on RNA replication. Pure Appl. Chem., 56, 967-978.
18. Matsumura, M. and Aiba, S. (1985) Screening for thermostable mutant of kanamycin nucleotidyltransferase by the use of a transformation system for a thermophile Bacillus stearothermophilus. J. Biol. Chem., 260, 15298-15303.
19. Bryan, P.N., Rollence, M.L., Pantoliano, M.W., Wood, J., Finzel, B.C., Gilliland, G.L., Howard, A.J., and Poulos, T.L. (1986) Proteases of enhanced stability: characterization of a thermostable variant of subtilisin. Proteins, 1, 326-334.
20. Leung, D.W., Chen, E., and Goeddel, D.V. (1989) A method for random mutagenesis of a defined DNA segment using a modified polymerase chain reaction. Technique, 1, 11-15;
21. Cadwell, R.C. and Joyce, G.F. (1994) Mutagenic PCR. PCR Methods Appl., 3, S136-S140.
22. Zhang, H.Y., Zhang, J., Lin, L., Du, W.Y., and Lu, J. (1993) Enhancement of the stability and activity of aspartase by random and site-directed mutagenesis. Biochem. Biophys. Res. Commun., 192, 15-21.
23. Liao, H., Mckenzie, T., and Hageman, R. (1986) Isolation of a thermostable enzyme variant by cloning and selection in a thermophile. Proc. Natl. Acad. Sci. U.S.A., 83, 576-580.
24. Chen, K. and Arnold, F.H. (1993) Tuning the activity of an enzyme for unusual environments: sequential random mutagenesis of subtilisin E for catalysis in dimethylformamide. Proc. Natl. Acad. Sci. U.S.A., 90, 5618-5622.
25. Stemmer, W.P.C. (1994) Rapid evolution of a protein in vitro by DNA shuffling. Nature, 370, 389-391.
26. Reetz, M.T., Zonta, A., Schimossek, K., Liebeton, K., and Jaeger, K.-E. (1997) Creation of enantioselective biocatalysts for organic chemistry by in vitro evolution. Angew. Chem., 109, 2961-2963; Angew. Chem. Int.Ed. Engl., 36, 2830-2832;
27. Reetz, M.T. (1999) Strategies for the development of enantioselective catalysts. Pure Appl. Chem., 71, 1503-1509.
28. Arnold, F.H. (1998) Design by directed evolution. Acc. Chem. Res., 31, 125-131.
29. Reetz, M.T. (2004) Controlling the enantioselectivity of enzymes by directed evolution: practical and theoretical ramifications. Proc. Natl. Acad. Sci. U.S.A., 101, 5716-5722.
30. Lutz, S. and Patrick, W.M. (2004) Novel methods for directed evolution of enzymes: quality, not quantity. Curr. Opin. Biotechnol., 15, 291-297.
31. Taylor, S.V., Kast, P., and Hilvert, D. (2001) Investigating and engineering enzymes by genetic selection. Angew. Chem., 113, 3408-3436; Angew. Chem. Int. Ed., 40, 3310-3335;
32. Lin, H. and Cornish, V.W. (2002) Screening and selection methods for large-scale analysis of protein function. Angew. Chem., 114, 4580-4606; Angew. Chem. Int. Ed., 41, 4402-4425;
33. Boersma, Y.L., Dröge, M.J., and Quax, W.J. (2007) Selection strategies for improved biocatalysts. FEBS J., 274, 2181-2195.
34. Reymond, J.-L. (2005) Enzyme Assays-High-Throughput Screening, Genetic Selection and Fingerprinting, Wiley-VCH VerlagGmbH, Weinheim;
35. Reetz, M.T. (2004) Screening for enantioselective enzymes, in Enzyme functionality: Design, Engineering and Screening (ed. A. Svendsen), Marcel Dekker, New York, pp. 559-598;
36. Reymond, J.-L., Fluxà, V.S., and Maillard, N. (2009) Enzyme assays. Chem. Commun., 34-46;
37. Polizzi, K.M., Parikh, M., Spencer, C.U., Matsumura, I., Lee, J.H., Realff, M.J., and Bommarius, A.S. (2006) Pooling for improved screening of combinatorial libraries for directed evolution. Biotechnol. Prog., 22, 961-967.
38. Selifonova, O. and Schellenberger, V. (2003) Evolution of microorganisms using mutator plasmids, in Directed Evolution Library Creation: Methods and Protocols (eds F.H. Arnold and G. Georgiou), Methods in Molecular Biology, vol. 231, Humana Press Inc., Totowa, NJ, pp. 45-52.
39. Wong, T.S., Roccatano, D., and Schwaneberg, U. (2007) Challenges of the genetic code for exploring sequence space in directed protein evolution. Biocatal. Biotransform., 25, 229-241.
40. Zaccolo, M., Williams, D.M., Brown, D.M., and Gherardi, E. (1996) An approach to random mutagenesis of DNA using mixtures of triphosphate derivatives of nucleoside analogous. J. Mol. Biol., 255, 589-603.
41. Claveau, S., Sasseville, M., and Beauregard, M. (2004) Alcohol-mediated error prone PCR. DNA Cell Biol., 23, 789-795.
42. Stratagene (2003) GeneMorph II Random Mutagenesis Kit: Instruction Manual, Stratagene, La Jolla, CA, U.S.A., http://www.bioc.rice.edu/bios576/proteng/200550.pdf
43. Eggert, T., Reetz, M.T., and Jaeger, K.-E. (2004) Directed evolution by random mutagenesis: a critical evaluation, in Enzyme Functionality-Design, Engineering, and Screening (ed. A. Svendsen), Marcel Dekker, New York, pp. 375-390;
44. Tee, K.L. and Schwaneberg, U. (2007) Directed evolution of oxygenases: screening system, success stories and challenges. Comb. Chem. High Throughput Screen., 10, 197-217.
45. Neylon, C. (2004) Chemical and biochemical strategies for the randomization of protein encoding DNA sequences: library construction methods for directed evolution. Nucleic Acids Res., 32, 1448-1459.
46. Sylvestre, J., Chautard, H., Cedrone, F., and Delcourt, M. (2006) Directed evolution of biocatalysts. Org. Process Res. Dev., 10, 562-571;
47. Cirino, P.C., Mayer, K.M. and Umeno, D. (2003) Generating mutant libraries using error-prone PCR, in Directed Evolution Library Creation: Methods and Protocols (eds F.H. Arnold and G. Georgiou), Humana Press, Totowa, NJ, pp. 3-10.
48. Fujii, R., Kitaoka, M., and Hayashi, K. (2006) Error-prone rolling circle amplification: the simplest random mutagenesis protocol. Nat. Protocols, 1, 2493-2497;
49. Fujii, R., Kitaoka, M., and Hayashi, K. (2004) One-step random mutagenesis by error-prone rolling circle amplification. Nucleic Acids Res., 32, e145.
50. Fromant, M., Blanquet, S., and Plateau, P. (1995) Direct random mutagenesis of gene-sized DNA fragments using polymerase chain reaction. Anal. Biochem., 224, 347-353.
51. Vanhercke, T., Ampe, C., Tirry, L., and Denolf, P. (2005) Reducing mutational bias in random protein libraries. Anal. Biochem., 339, 9-14.
52. Wong, T.S., Tee, K.L., Hauer, B., and Schwaneberg, U. (2004) Sequence saturation mutagenesis (SeSaM): a novel method for directed evolution. Nucleic Acids Res., 32, e26.
53. Wong, T.S., Roccatano, D., Loakes, D., Tee, K.L., Schenk, A., Hauer, B., and Schwaneberg, U. (2008) Transversionenriched sequence saturation mutagenesis (SeSaM-Tv+): a random mutagenesis method with consecutive nucleotide exchanges that complements the bias of error-prone PCR. Biotechnol. J., 3, 74-82
54. Shivange, A.V., Marienhagen, J., Mundhada, H., Schenk, A. and Schwaneberg, U. (2009) Advances in generating functional diversity for directed protein evolution. Curr. Opin. Chem. Biol., 13, 19-25.
55. Murakami, H., Hohsaka, T., and Sisido, M. (2002) Random insertion and deletion of arbitrary number of bases for codonbased random mutation of DNAs. Nat. Biotechnol., 20, 76-81.
56. Pikkemaat, M.G. and Janssen, D.B. (2002) Generating segmental mutations in haloalkane dehalogenase: a novel partin the directed evolution toolbox. Nucleic Acids Res., 30, e35;
57. Jones, D.D. (2005) Triplet nucleotide removal at random positions in a target gene: the tolerance of TEM-1 b-lactamase to an amino acid deletion. Nucleic Acids Res., 33, e80.
58. Erdogan, E., Jones, R.J., Matzlin, P., Hanna, M.H., Smith, S.M.E., and Salerno, J.C. (2005) A novel mutagenesis method generating high yields of closed circularmutant DNAwith one primer per mutant. Mol. Biotechnol., 30, 21-30.
59. Fujii, R., Kitaoka, M., and Hayashi, K. (2006) RAISE: a simple and novel method of generating random insertion and deletion mutations. Nucleic Acids Res., 34, e30.
60. Hogrefe, H.H., Cline, J., Youngblood, G.L., and Allen, R.M. (2002) Creating randomized amino acid libraries with the QuikChange multi site-directed mutagenesis kit. BioTechniques, 33, 1158-1165.
61. Dube, D.K. and Loeb, L.A. (1989) Mutants generated by the insertion of random oligonucleotides into the active site of the beta-lactamase gene. Biochemistry, 28, 5703-5707;
62. Teplyakov, A.V., Van Der Laan, J.M., Lammers, A.A., Kelders, H., Kalk, K.H., Misset, O., Mulleners, L.J.S.M., and Dijkstra, B.W. (1992) Protein engineering of the high-alkaline serine protease PB92 from Bacillus alcalophilus: functional and structural consequences of mutation at the S4 substrate binding pocket. Protein Eng., 5, 413-420;
63. Graham, L.D., Haggett, K.D., Jennings, P.A., Le Brocque, D.S., Whittaker, R.G., and Schober, P.A. (1993) Random mutagenesis of the substrate-binding site of a serine protease can generate enzymes with increased activities and altered primary specificities. Biochemistry, 32, 6250-6258;
64. Warren, M.S. and Benkovic, S.J. (1997) Combinatorial manipulation of three key active site residues in glycinamide ribonucleotide transformylase. Protein Eng., 10, 63-68.
65. Reetz, M.T. and Carballeira, J.D. (2007) Iterative saturation mutagenesis (ISM) for rapid directed evolution of functional enzymes. Nat. Protocols, 2, 891-903;
66. CASTER and B-FITTER are available free of charge on the author's homepage (http://www.kofo.mpg.de/en/research/organic-synthesis)
67. Firth, A.E. and Patrick, W.M. (2005) Statistics of protein library construction. Bioinformatics, 21, 3314-3315.
68. Firth, A.E. and Patrick, W.M. (2008) GLUE-IT and PEDEL-AA: new programmes for analyzing protein diversity in randomized libraries. Nucleic Acids Res., 36, W281-W285.
69. Hughes, M.D., Nagel, D.A., Santos, A.F., Sutherland, A.J., and Hine, A.V. (2003) Removing the redundancy from randomised gene libraries. J. Mol. Biol., 331, 973-979.
70. Evans, P.M. and Liu, C. (2005) SiteFind: a software tool for introducing a restriction site as a marker for successful sitedirected mutagenesis. BMC Mol. Biol., 6, 22.
71. Miyazaki, K. and Takenouchi, M. (2002) Creating random mutagenesis libraries using megaprimer PCR of whole plasmid. BioTechniques, 33, 1033-1038.
72. Sanchis, J., Fernández, L., Carballeira, J.D., Drone, J., Gumulya, Y., Höbenreich, H., Kahakeaw, D., Kille, S., Lohmer, R., Peyralans, J.J.-P., Podtetenieff, J., Prasad, S., Soni, P., Taglieber, A., Wu, S., Zilly, F.E., and Reetz, M.T. (2008) Improved PCR method for the creation of saturation mutagenesis libraries in directed evolution: application to difficult-to-amplify templates. Appl. Microbiol. Biotechnol., 81, 387-397.
73. Kirsch, R.D. and Joly, E. (1998) An improved PCR-mutagenesis strategy for two-site mutagenesis or sequence swapping between related genes. Nucleic Acids Res., 26, 1848-1850;
74. Zheng, L., Baumann, U., and Reymond, J.-L. (2004) An efficient onestep site-directed and site-saturation mutagenesis protocol. Nucleic Acids Res., 32, e115.
75. Rui, L., Kwon, Y.M., Fishman, A., Reardon, K.F., and Wood, T.K. (2004) Saturation mutagenesis of toluene orthomonooxygenase of Burkholderia cepacia G4 for enhanced 1-naphthol synthesis and chloroform degradation. Appl. Environ. Microbiol., 70, 3246-3252.
76. Reetz, M.T., Kahakeaw, D. and Lohmer, R. (2008) Addressing the numbers problem in directed evolution. ChemBioChem, 9, 1797-1804;
77. Reetz, M.T., Kahakeaw, D., and Sanchis, J. (2009) Shedding light on the efficacy of laboratory evolution based on iterative saturation mutagenesis. Molecular BioSystems, 5, 115-122.
78. Patrick, W.M. and Firth, A.E. (2005) Strategies and computational tools for improving randomized protein libraries. Biomol. Eng., 22, 105-112;
79. Bosley, A.D. and Ostermeier, M. (2005) Mathematical expressions useful in the construction, description and evaluation of protein libraries. Biomol. Eng., 22, 57-61;
80. Denault, M. and Pelletier, J.N. (2007) Protein Library Design and Screening: Working Out the Probabilities, in Protein Engineering Protocols, vol. 352 (eds K.M. Arndt and K.M. Müller), Humana Press, Totowa, NJ, pp. 127-154.
81. Iyidogan, P. and Lutz, S. (2008) Systematic exploration of active site mutations on human deoxycytidine kinase substrate specificity. Biochemistry, 47, 4711-4720.
82. Van den Brulle, J., Fischer, M., Langmann, T., Horn, G., Waldmann, T., Arnold, S., Fuhrmann, M., Schatz, O., O'Connell, T., O'Connell, D., Auckenthaler, A., and Schwer, H. (2008) A novel solid phase technology for highthroughput gene synthesis. BioTechniques, 45, 340-343.
83. Clouthier, C.M., Kayser, M.M., and Reetz, M.T. (2006) Designing new Baeyer-Villiger monooxygenases using restricted CASTing. J. Org. Chem., 71, 8431-8437.
84. Bougioukou, D.J., Kille, S., Taglieber, A., and Reetz, M.T. (2009) Directed evolution of an enantioselective enoate-reductase: Testing the utility of iterative saturation mutagenesis. Adv. Synth. Catal., 351, 3287-3305.
85. Climie, S., Ruiz-Perez, L., Gonzalez-Pacanowska, D., Prapunwattana, P., Cho, S.-W., Stroud, R. and Santi, D.V. (1990) Saturation site-directed mutagenesis of thymidylate synthase. J. Biol. Chem., 265, 18776-18779;
86. Gabor, E.M. and Janssen, D.B. (2004) Increasing the synthetic performance of penicillin acylase PAS2 by structure-inspired semirandom mutagenesis. Protein Eng., Design & Select., 17, 571-579;
87. Rui, L., Cao, L., Chen, W., Reardon, K.F. and Wood, T.K. (2004) Active site engineering of the epoxide hydrolase from Agrobacterium radiobacter AD1 to enhance aerobic mineralization of cis-1,2-dichloroethylene in cells expressing an evolved toluene orthomonooxygenase. J. Biol. Chem., 279, 46810-46817;
88. Schmitzer, A.R., Lepine, F., and Pelletier, J.N. (2004) Combinatorial exploration of the catalytic site of a drug-resistant dihydrofolate reductase: creating alternative functional configurations. Protein Eng., Design & Select., 17, 809-819;
89. Antikainen, N.M., Hergenrother, P.J., Harris, M.M., Corbett, W. and Martin, S.F. (2003) Altering substrate specificity of phosphatidylcholine-preferring phospholipase c of Bacillus cereus by random mutagenesis of the headgroup binding site. Biochemistry, 42, 1603-1610.
90. Reetz, M.T., Wilensek, S., Zha, D., and Jaeger, K.-E. (2001) Directed evolution of an enantioselective enzyme through combinatorial multiple cassette mutagenesis. Angew. Chem., 113, 3701-3703; Angew. Chem. Int. Ed., 40, 3589-3591.
91. Horsman, G.P., Liu, A.M.F., Henke, E., Bornscheuer, U.T. and Kazlauskas, R.J. (2003) Mutations indistant residues moderately increase the enantioselectivity of Pseudomonas fluorescens esterase towards methyl 3-bromo-2-methylpropanoate and ethyl 3-phenylbutyrate. Chem. Eur. J., 9, 1933-1939;
92. Park, K.L., Morley, K.L., Horsman, G.P., Holmquist, M., Hult, K., and Kazlauskas, R.J. (2005) Focusing mutations into the P. fluorescens esterase binding site increases enantioselectivity more effectively than distant mutations. Chem. Biol., 12, 45-54;
93. Kazlauskas, R.J. (2003) Creating enantioselective hydrolases for organic synthesis: combining rational and randommethods. Chem. Listy, 97, 329-337.
94. Paramesvaran, J., Hibbert, E.G., Russell, A.J., and Dalby, P.A. (2009) Distributions of enzyme residues yielding mutants with improved substrate specificities from two different directed evolution strategies. Protein Eng., Design & Select., 22, 401-411.
95. Reetz, M.T., Wang, L.-W., and in part Bocola, M. (2006) Directed evolution of enantioselective enzymes: iterative cycles of CASTing for probing proteinsequence space. Angew. Chem., 118, 1258-1263, erratum, p. 2556; Angew. Chem. Int. Ed. Engl., 45, 1236-1241, erratum, p. 2494.
96. Reetz, M.T., Prasad, S., Carballeira, J.D., Gumulya, Y., and Bocola, M. (2010) Iterative saturation mutagenesis accelerates laboratory evolution of enzyme stereoselectivity: rigorous comparison with traditional approaches. J. Am. Chem. Soc., 132, 9144-9152;
97. Gumulya, Y. and Reetz, M.T. (2011) Enhancing the thermal robustness of an enzyme by directed evolution: Least favorable starting points and inferior mutants can map superior evolutionary pathways. ChemBioChem., 12, 2502-2510.
98. Reetz, M.T., Carballeira, J.D., and Vogel, A. (2006) Iterative saturation mutagenesis on the basis of B factors as a strategy for increasing protein thermostability. Angew. Chem., 118, 7909-7915; Angew. Chem. Int. Ed., 45, 7745-7751.
99. Reetz, M.T., Bocola, M., Carballeira, J.D., Zha, D., and Vogel, A. (2005) Expanding the range of substrate acceptance of enzymes: combinatorial active-site saturation test. Angew. Chem., 117, 4264-4268; Angew. Chem. Int. Ed., 44, 4192-4196.
100. Oshima, T. (1994) Stabilization of proteins by evolutionary molecular engineering techniques. Curr. Opin.Struct. Biol., 4, 623-628;
101. Ó'Fágáin, C. (2003) Enzyme stabilization-recent experimental progress. Enzyme Microb. Technol., 33, 137-149;
102. Eijsink, V.G.H., Gaseidnes, S., Borchert, T.V., and Van Den Burg, B. (2005) Directed evolution of enzyme stability. Biomol. Eng., 22, 21-30;
103. Bommarius, A.S. and Broering, J.M. (2005) Established and novel tools to investigate biocatalyst stability. Biocatal. Biotransform., 23, 125-139.
104. Chaparro-Riggers, J.F., Polizzi, K.M., and Bommarius, A.S. (2007) Better library design: data-driven protein engineering. Biotechnol. J., 2, 180-191;
105. Gustafsson, C., Govindarajan, S., and Minshull, J. (2003) Putting engineering back into protein engineering: bioinformatic approaches to catalyst design. Curr. Opin. Biotechnol., 14, 366-370.
106. Steipe, B., Schiller, B., Plückthun, A., and Steinbacher, S. (1994) Sequence statistics reliably predict stabilizing mutations in a protein domain. J. Mol. Biol., 240, 188-192;
107. Dai, M., Fisher, H.E., Temirov, J., Kiss, C., Phipps, M.E., Pavlik, P., Werner, J.H., and Bradbury, A.R.M. (2007) The creation of a novel fluorescent protein by guided consensus engineering. Protein Eng., Design & Select., 20, 69-79;
108. Amin, N., Liu, A.D., Ramer, S., Aihle, W., Meijer, D., Metin, M., Wong, S., Gualfetti, P., and Schellenberger, V. (2004) Construction of stabilized proteins by combinatorial consensus mutagenesis. Protein Eng., Design & Select., 17, 787-793;
109. Hamamatsu, N., Aita, T., Nomiya, Y., Uchiyama, H., Nakajima, M., Husimi, Y., and Shibanaka, Y. (2005) Biased mutation-assembling: an efficient method for rapid directed evolution through simultaneous mutation accumulation. Protein Eng., Design & Select., 18, 265-271.
110. Rungpragayphan, S., Kawarasaki, Y., Imaeda, T., Kohda, K., Nakano, H., and Yamane, T. (2002) High-throughput, cloning-independent protein library construction by combining singlemolecule DNA amplification with in vitro expression. J. Mol. Biol., 318, 395-405;
111. Koga, Y., Kato, K., Nakano, H., and Yamane, T. (2003) Inverting enantioselectivity of Burkholderia cepacia KWI-56 lipase by combinatorial mutation and high-throughput screening using single-molecule PCR and in vitro expression. J. Mol. Biol., 331, 585-592.
112. Joern, J.M. (2003) DNA shuffling, in Directed Evolution Library Creation (eds F.H. Arnold and G. Georgiou), Humana Press Inc., Totowa, NJ, pp. 85-89.
113. Powell, K.A., Ramer, S.W., del Cardayre, S.B., Stemmer, W.P.C., Tobin, B., Longchamp, P.F., and Huismann, G.W. (2001) Directed evolutionand biocatalysis. Angew. Chem., 113, 4068-4080; Angew. Chem. Int. Ed., 44, 3948-3959.
114. Joern, J.M., Meinhold, P., and Arnold, F.H. (2002) Analysis of shuffled gene libraries. J. Mol. Biol., 316, 643-656.
115. Zhao, H., Giver, L., Shao, Z., Affholter, J.A., and Arnold, F.H. (1998) Molecular evolution by staggered extension process (StEP) in vitro recombination. Nat. Biotechnol., 16, 258-261.
116. Ostermeier, M., Shim, J.H., and Benkovic, S.J. (1999) A combinatorial approach to hybrid enzymes independent of DNA homology. Nat. Biotechnol., 17, 1205-1209;
117. Lutz, S., Ostermeier, M., and Benkovic, S.J. (2001) Rapid generation of incremental truncation libraries for protein engineering using a-phosphothioate nucleotides. Nucleic Acids Res., 29, e16.
118. Kawarasaki, Y., Griswold, K.E., Stevenson, J.D., Selzer, T., Benkovic, S.J., Iverson, B.L., and Georgiou, G. (2003) Enhanced crossover SCRATCHY: construction and high-throughput screening of a combinatorial library containing multiple non-homologous crossovers. Nucleic Acids Res., 31, e126.
119. Sieber, V., Martinez, C.A., and Arnold, F.H. (2001) Libraries of hybrid proteins from distantly related sequences. Nat. Biotechnol., 19, 456-460.
120. Higara, K. and Arnold, F.H. (2003) General method for sequenceindependent site-directed chimeragenesis. J. Mol. Biol., 330, 287-296.
121. Lee, S.H., Ryu, E.J., Kang, M.J., Wang, E.-S., Piao, Z., Choi, Y.J., Jung, K.H., Jeon, J.Y.J., and Shin, Y.C. (2003) A new approach to directed gene evolution by recombined extension on truncated templates (RETT). J. Mol. Catal., B: Enzym., 26, 119-129.
122. Ikeuchi, A., Kawarasaki, Y., Shinbata, T., and Yamane, T. (2003) Chimeric gene library construction by a simple and highly versatile method using recombination-dependent exponential amplification. Biotechnol. Prog., 19, 1460-1467.
123. O'Maille, P.E., Bakhtina, M., and Tsai, M.D. (2002) Structure-based combinatorial protein engineering (SCOPE). J. Mol. Biol., 321, 677-691.
124. Coco, W.M., Levinson, W.E., Crist, M.J., Hektor, H.J., Darzins, A., Pienkos, P.T., Squires, C.H., and Monticello, D.J. (2001) DNA shuffling method for generating highly recombined genes and evolved enzymes. Nat. Biotechnol., 19, 354-359.
125. Gibbs, M.D., Nevalainen, K.M.H., and Bergquist, P.L. (2001) Degenerate oligonucleotide gene shuffling (DOGS): a method for enhancing the frequency of recombination with family shuffling. Gene, 271, 13-20.
126. Bergquist, P.L., Reeves, R.A., and Gibbs, M.D. (2005) Degenerate oligonucleotide gene shuffling (DOGS) and random drift mutagenesis (RNDM): two complementary techniques for enzyme evolution. Biomol. Eng., 22, 63-72.
127. Reiter, B., Faschinger, A., Glieder, A., and Schwab, H. (2007) Random strand transfer recombination (RSTR) for homology-independent nucleic acid recombination. J. Biotechnol., 129, 39-49.
128. Stemmer, W.P.C. (1994) DNA shuffling by random fragmentation and reassembly: in vitro recombination for molecular evolution. Proc. Natl. Acad. Sci. U.S.A., 91, 10747-10751;
129. Stutzman-Engwall, K., Conlon, S., Fedechko, R., Mcarthur, H., Pekrun, K., Chen, Y., Jenne, S., La, C., Trinh, N., Kim, S., Zhang, Y.-X., Fox, R., Gustafsson, C., and Krebber, A. (2005) Semi-synthetic DNA shuffling of aveC leads to improved industrial scale production of doramectin by Streptomyces avermitilis.Metab. Eng., 7, 27-37.
130. Herman, A. and Tawfik, D.S. (2007) Incorporating synthetic oligonucleotides via gene reassembly (ISOR): a versatile tool for generating targeted libraries. Protein Eng., Design & Select., 20, 219-226.
131. Coco, W.M., Encell, L.P., Levinson, W.E., Crist, M.J., Loomis, A.K., Licato, L.L., Arensdorf, J.J., Sica, N., Pienkos, P.T., and Monticello, D.J. (2002)Growth factor engineering by degenerate homoduplex gene family recombination. Nat. Biotechnol., 20, 1246-1250.
132. Ness, J.E., Kim, S., Gottman, A., Pak, R., Krebber, A., Borchert, T.V., Govindarajan, S., Mundorff, E.C., and Minshull, J. (2002) Synthetic shuffling expands functional protein diversity by allowing amino acids to recombine independently. Nat. Biotechnol., 20, 1251-1255.
133. Zha, D., Eipper, A., and Reetz, M.T. (2003) Assembly of designed oligonucleotides as an efficient method for gene recombination: a new tool in directed evolution. ChemBioChem, 4, 34-39.
134. Li, Y. and Palmer, A.G. III (2009) Domain swapping in the kinase superfamily: OSR1 joins the mix. Protein Sci., 18, 678-681.
135. Park, S.-H., Park, H.-Y., Sohng, J.K., Lee, H.C., Liou, K., Yoon, Y.J., and Kim, B.-G. (2009) Expanding substrate specificity of gt-b fold glycosyltransferase via domain swapping and highthroughput screening. Biotechnol. Bioeng., 102, 988-994;
136. Cartwright, A.M., Lim, E.-K., Kleanthous, C., and Bowles, D.J. (2008) A kinetic analysis of regiospecific glucosylation by two glycosyltransferases of Arabidopsis thaliana. J. Biol. Chem., 282, 15724-15731;
137. Hansen, E.H., Osmani, S.A., Kristensen, C., Møller, B.L., and Hansen, J. (2009) Substrate specificities of family 1 UGTs gained by domain swapping. Phytochemistry, 70, 473-482.
138. Sharkey, M.A. and Engel, P.C. (2009) Modular coenzyme specificity; a domainswopped chimera of glutamate dehydrogenase. Proteins, 77, 268-278.
139. Meister, G.E., Kanwar, M., and Ostermeier, M. (2009) Circular permutation pf proteins, in Protein Engineering Handbook, vol. 2 (eds S. Lutz and U.T. Bornscheuer), Wiley-VCH Verlag GmbH, Weinheim, pp. 454-471.
140. Qian, Z. and Lutz, S. (2005) Improving the catalytic activity of Candida antarctica lipase b by circular permutation. J. Am. Chem. Soc., 127, 13466-13467;
141. Yu, Y. and Lutz, S. (2010) Improved triglyceride transesterification by circular permuted Candida antarctica lipase B. Biotechnol. Bioeng., 105, 44-50.
142. Reitinger, S., Yu, Y., Wicki, J., Ludwiczek, M., D'Angelo, I., Baturin, S., Okon, M., Strynadka, N.C.J., Lutz, S., Withers, S.G., and Mcintosh, L.P. (2010) Circular permutation of Bacillus circulans xylanase: a kinetic and structural study. Biochemistry. 49 (11), 2464-2474.
143. Lo, W.-C. and Lyu, P.-C. (2008) CPSARST: an efficient circular permutation search tool applied to the detection of novel protein structural relationships. GenomeBiology, 9, R11.
144. Drummond, D.A., Iverson, B.L., Georgiou, G., and Arnold, F.H. (2005) Why high-error-rate random mutagenesis libraries are enriched in functional and improved proteins. J. Mol. Biol., 350, 806-816.
145. Tracewell, C.A. and Arnold, F.H. (2009) Directed enzyme evolution: climbing fitness peaks one amino acid at a time. Curr. Opin. Chem. Biol., 13, 3-9.
146. Bloom, J.D., Labthavikul, S.T., Otey, C.R., and Arnold, F.H. (2006) Protein stability promotes evolvability. Proc. Natl. Acad. Sci. U.S.A., 103, 5869-5874.
147. Peisajovich, S.G. and Tawfik, D.S. (2007) Protein engineers turned evolutionists. Nat. Methods, 4, 991-994;
148. Bloom, J.D. and Arnold, F.H. (2009) In the light of directed evolution: pathways of adaptive protein evolution. Proc. Natl. Acad. Sci. U.S.A., 106, 9995-10000;
149. Depristo, M.A. (2007) The subtle benefits of being promiscuous: adaptive evolution potentiated by enzyme promiscuity. HFSP J., 1, 94-98.;
150. Kryazhimskiy, S., Dushof, J., Bazykin, G.A., and Plotkin, J. B. (2011) Prevalence of epistasis in the evolution of influenza A surface proteins. PLoS Genetics, 7, e1001301.
151. Eigen, M., McCaskill, J., and Schuster, P. (1988) Molecular quasi-species. J. Phys. Chem., 92, 6881-6891.
152. Kurtovic, S. and Mannervik, B. (2009) Identification of emerging quasi-species in directed enzyme evolution, Biochemistry, 48, 9330-9339.
153. Runarsdottir, A. and Mannervik, B. (2010) A novel quasi-species of glutathione transferase with high activity towards naturally occurring isothiocyanates evolves from promiscuous low-activity variants. J. Mol. Biol., 401, 451-464.
154. Parikh, M.R. and Matsumura, I. (2005) Site-saturation mutagenesis is more efficient than DNA shuffling for the directed evolution of β-fucosidase from b-galactosidase. J. Mol. Biol., 352, 621-628.
155. Pavlova, M., Klvana, M., Prokop, Z., Chaloupkova, R., Banas, P., Otyepka, M., Wade, R.C., Tsuda, M., Nagata, Y., and Damborsky, J. (2009) Redesigning dehalogenase access tunnels as a strategy for degrading an anthropogenic substrate. Nat. Chem. Biol., 5, 727-733.
156. Wang, T.-W., Zhu, H., Ma, X.-Y., Zhang, T., Ma, Y.-S., andWei, D.-Z. (2006) Mutant library construction in directed molecular evolution. Mol. Biotechnol., 34, 55-68.
157. Zhang, J.-H., Dawes, G., and Stemmer, W.P.C. (1997) Directed evolution of a fucosidase from a galactosidase by DNA shuffling and screening. Proc. Natl. Acad. Sci. U.S.A., 94, 4504-4509.
158. Juers, D.H., Heightman, T.D., Vasella, A., McCarter, J.D., Mackenzie, L., Withers, S.G., and Matthews, B.W. (2001) A structural view of the action of Escherichia coli (lacZ) β-galactosidase. Biochemistry, 40, 14781-14794.
159. Reetz, M.T., Torre, C., Eipper, A., Lohmer, R., Hermes, M., Brunner, B., Maichele, A., Bocola, M., Arand, M., Cronin, A., Genzel, Y., Archelas, A., and Furstoss, R. (2004) Enhancing the enantioselectivity of an epoxide hydrolase by directed evolution. Org. Lett., 6, 177-180.
160. Reetz, M.T. and Sanchis, J. (2008) Constructing and analyzing the fitness landscape of an experimental evolutionary process. ChemBioChem, 9, 2260-2267.
161. Liebeton, K., Zonta, A., Schimossek, K., Nardini, M., Lang, D., Dijkstra, B.W., Reetz, M.T., and Jaeger, K.-E. (2000) Directed evolution of an enantioselective lipase. Chem. Biol., 7, 709-718.
162. Nardini, M., Lang, D.A., Liebeton, K., Jaeger, K.-E., and Dijkstra, B.W. (2000) Crystal structure of Pseudomonas aeruginosa lipase in the open conformation. J. Biol. Chem., 275, 31219-31225.
163. Hughes, M.D., Zhang, Z.-R., Sutherland, A.J., Santos, A.F., and Hine, A.V. (2005) Discovery of active proteins directly from combinatorial randomized protein libraries without display, purification or sequencing: identification of novel zinc finger proteins. Nucleic Acids Res., 33, e32;
164. Chockalingam, K., Chen, Z., Katzenellenbogen, J.A., and Zhao, H. (2005) Directed evolution of specific receptor-ligand pairs for use in the creation of gene switches. Proc. Natl. Acad. Sci. U.S.A., 102, 5691-5696.
165. Crameri, A., and Stemmer, W.P.C. (1995) Combinatorial multiple cassette mutagenesis creates all the permutations of mutant and wild-type sequences. BioTechniques, 18, 194-196.
166. Reetz, M.T., Puls, M., Carballeira, J.D., Vogel, A., Jaeger, K.-E., Eggert, T., Thiel, W., Bocola, M., and Otte, N. (2007) Learning from directed evolution: further lessons from theoretical investigations into cooperative mutations in lipase enantioselectivity. ChemBioChem, 8, 106-112.
167. Sandström, A.G., Engström, K., Nyhlen, J., Kasrayan, A. and Bäckvall, J.-E. (2009) Directed evolution of Candida antarctica lipase A using an episomaly replicating yeast plasmid. Protein Eng., Design & Select., 22, 413-420;
168. Engström, K., Nyhlen, J., Sandtröm, A.G., and Bäckvall, J.-E. (2010) Directed evolution of an enantioselective lipase with broad substrate scope for hydrolysis of αsubstituted esters. J. Am. Chem. Soc., 132, 7038-7042.
169. Okrasa, K., Levy, C., Wilding, M., Goodall, M., Baudendistel, N., Hauer, B., Leys, D., and Micklefield, J. (2009) Structureguided directed evolution of alkenyl and arylmalonate decarboxylases. Angew. Chem., 121, 7827-7830; Angew. Chem. Int. Ed., 48, 7691-7694.
170. Hawwa, R., Larsen, S.D., Ratia, K., and Mesecar, A.D. (2009) Structure-based and random mutagenesis approaches increase the organophosphate-degrading activity of a phosphotriesterase homologue from Deinococcus radiodurans. J. Mol. Biol., 393, 36-57.
171. Ivancic, M., Valinger, G., Gruber, K., and Schwab, H. (2007) Inverting enantioselectivity of Burkholderia gladioli esterase EstB by directed and designed evolution. J. Biotechnol., 129, 109-122.
172. De Groeve, M.R.M., De Baere, M., Hoflack, L., Desmet, T., Vandamme, E.J., and Soetaert, W. (2009) Creating lactose phosphorylase enzymes by directed evolution of cellobiose phosphorylase. Protein Eng., Design & Select., 22, 393-399.
173. Liang, L., Zhang, J., and Lin, Z. (2007) Altering coenzyme specificity of Pichia stipitis xylose reductase by the semirational approachCASTing. MicrobialCell Factories, 6, 36.
174. Kelly, R.M., Leemhuis, H., and Dijkhuizen, L. (2007) Conversion of a cyclodextrin glucanotransferase into an αamylase: assessment of directed evolution strategies. Biochemistry, 46, 11216-11222.
175. Bhuiya M.-W. and Liu C.-J. (2010) engineering monolignol 4-Omethyltransferase to modulate lignin biosynthesis. J. Biol. Chem., 285, 277-313.
176. Reetz, M.T., Peyralans, J.J.-P., Maichele, A., Fu, Y., and Maywald, M. (2006) Directed evolution of hybrid enzymes: evolving enantioselectivity of an achiral Rhcomplex anchored to a protein. Chem. Commun., 4318-4320.
177. Pavelka, A., Chovancova, E., and Damborsky, J. (2009) HotSpot Wizard: a web-server for identification of hot spots in protein engineering. Nucleic Acids Res., 37, W376-W383.
178. Saven, J.G. (2009) Computational protein design, in Protein Engineering Handbook, vol. 1 (eds S. Lutz and U.T. Bornscheuer), Wiley-VCH Verlag GmbH, Weinheim, pp. 325-342.
179. Damborsky, J. and Brezovsky, J. (2009) Computational tools for designing end engineering biocatalysts. Curr. Opin. Chem. Biol., 13, 26-34;
180. Braiuca, P., Lorena, K., Ferrario, V., Ebert, C., and Gardossi, L. (2009) A three-dimensional quantitative structure-activity relationship (3D-QSAR) model for predicting the enantioselectivity of Candida antarctica lipase B. Adv. Synth. Catal., 351, 1293-1302;
181. Martí, S., Andrés, J., Moliner, V., Silla, E., and Tuñón, I. (2008) Computational design of biological catalysts. Chem. Soc. Rev., 37, 2634-2643;
182. Sterner, R., Merkl, R., and Raushel, F.M. (2008) Computational design of enzymes. Chem. Biol., 15, 421-423;
183. Chiappori, F., D'ursi, P., Merelli, I., Milanesi, L. and Rovida, E. (2009) In silico saturation mutagenesis and docking screening for the analysis of protein-ligand interaction: the endothelial protein C receptor case study. BMC Bioinformatics, 10, S3;
184. Senn, H.M. and Thiel, W. (2009) QM/MM methods for biomolecular systems. Angew. Chem., 121, 1220-1254; Angew. Chem. Int. Ed., 48, 1198-1229 ;
185. Siegel, J.B., Zanghellini, A., Lovick, H.M., Kiss, G., Lambert, A.R., St.Clair, J.L., Gallaher, J.L., Hilvert, D., Gelb, M.H., Stoddard, B.L., Houk, K.N., Michael, F.E., and Baker, D. (2010) Computational design of an enzyme catalyst for a stereoselective biomolecular Diels-Alder reaction. Science, 329, 309-313.
186. Reetz, M.T. and Wu, S. (2008) Greatly reduced amino acid alphabets in directed evolution: making the right choice for saturation at homologous enzyme positions. Chem. Commun., 5499-5501.
187. Saab-Rincon, G., Li, Y., Meyer, M., Carbone, M., Landwehr, M., and Arnold, F.H. (2009) Protein engineering by structure-guided SCHEMA recombination, in Protein Engineering Handbook (eds S. Lutz and U.T. Bornscheuer), Wiley-VCH Verlag GmbH, Weinheim, pp. 481-492
188. Heinzelman, P., Snow, C.D., Smith, M.A., Yu, X., Kannan, A., Boulware, K., Villalobos, A., Govindarajan, S., Minshull, J., and Arnold, F.H. (2009) SCHEMA recombination of a fungal cellulase uncovers a single mutation that contributes markedly to stability. J. Biol. Chem., 284, 26229-26233.
189. Hernández, G. and Lemaster, D.M. (2005) Hybrid native partitioning of interactions among nonconserved residues in chimeric proteins. Proteins, 60, 723-731.
190. Saraf, M.C., Horswill, A.R., Benkovic, S.J., andMaranas, C.D. (2004) FamClash: a method for ranking the activity of engineered enzymes. Proc. Natl. Acad. Sci. U.S.A., 101, 4142-4147.
191. Pantazes, R.J., Saraf, M.C., and Maranas, C.D. (2007) Optimal protein library design using recombination or point mutations based on sequencebased scoring functions. Protein Eng., Design Select., 20, 361-373.
192. Wedge, D.C., Rowe, W., Kell, D.B., and Knowles, J. (2009) In silico modelling of directed evolution: implications for experimental design and stepwise evolution. J. Theor. Biol., 257, 131-141.
193. Fox, R.J., Davis, S.C., Mundorff, E.C., Newman, L.M., Gavrilovic, V., Ma, S.K., Chung, L.M., Ching, C., Tam, S., Muley, S., Grate, J., Gruber, J., Whitman, J.C., Sheldon, R.A., and Huisman, G.W. (2007) Improving catalytic function by ProSAR-driven enzyme evolution. Nat. Biotechnol., 25, 338-344.
194. Liang, J., Lalonde, J., Borup, B., Mitchell, V., Mundorff, E., Trinh, N., Kochrekar, D.A., Cherat, R.N., and Pai, G.G. (2010) Development of a biocatalytic process as an alternative to the (-)-DIP-CI-mediated asymmetric reduction of a key intermediate of montelukast. Org. Process Res. Dev., 14, 193-198.
195. Saraf, M.C., Moore, G.L., Goodey, N.M., Cao, V.Y., and Benkovic, S.J. (2006) IPRO: an iterative computational protein library redesign and optimization procedure. Biophys. J., 90, 4167-4180.
196. Chen, R., Li, L., and Weng, Z. (2003) ZDOCK: an initial-stage protein-docking algorithm. Proteins, 52, 80-87.
197. Khoury, G.A., Fazelinia, H., Chin, J.W., Pantazes, R.J., Cirino, P.C., and Maranas, C.D. (2009) Computational design of Candida boidinii xylose reductase for altered cofactor specificity. Protein Sci., 18, 2125-2138.
198. Fazelinia, H., Cirino, P.C., and Maranas, C.D. (2009) OptGraft: a computational procedure for transferring a binding site onto an existing protein scaffold. Protein Sci., 18, 180-195.
199. Reetz, M.T., Kühling, K.M., Wilensek, S., Husmann, H., Häusig, U.W., and Hermes, M. (2001) A GC-based method for high-throughput screening of enantioselective catalysts. Catal. Today, 67, 389-396;
200. Reetz, M.T., Daligault, F., Brunner, B., Hinrichs, H., and Deege, A. (2004) Directed evolution of cyclohexanone monooxygenases: enantioselective biocatalysts for the oxidation of prochiral thioethers. Angew. Chem., 116, 4170-4173; Angew. Chem. Int. Ed., 43, 4078-4081;
201. Reetz, M.T., Becker, M.H., Klein, H.-W., and Stöckigt, D. (1999) A method for highthroughput screening of enantioselective catalysts. Angew. Chem., 111, 1872-1875; Angew. Chem. Int. Ed., 38, 1758-1761.
202. Trapp, O. (2007) Boosting the throughput of separation techniques by "multiplexing". Angew. Chem., 119, 5706-5710; Angew. Chem. Int. Ed., 46, 5609-5613.
203. Gumulya, Y. (2010) Directed evolution of enantioselective epoxide hydrolases, Dissertation, Ruhr-Universität Bochum, 2010.
204. Reetz, M.T. (2001) Combinatorial and evolution-based methods in the creation of enantioselective catalysts. Angew. Chem., 113, 292-320; Angew. Chem. Int. Ed., 40, 284-310.
205. Boersma, Y.L., Dröge, M.J., Van Der Sloot, A.M., Pijning, T., Cool, R.H., Dijkstra, B.W., and Quax, W.J. (2008) A novel genetic selection system for improved enantioselectivity of Bacillus subtilis lipase A. ChemBioChem, 9, 1110-1115.
206. Reetz, M.T., Höbenreich, H., Soni, P., and Fernández, L. (2008) A genetic selection system for evolving enantioselectivity of enzymes. Chem. Commun., 5502-5504.
207. Griffiths, A.D. and Tawfik, D.S. (2006) Miniaturising the laboratory in emulsion droplets. Trends in Biotechnol., 24, 395-402;
208. Aharoni, A. and Tawfik, D.S. (2009) In vitro compartmentalization (IVC) and other high-throughput screens of enzyme libraries, in Protein Engineering Handbook, vol. 2 (eds S. Lutz and U.T. Bornscheuer), Wiley-VCH Verlag GmbH, Weinheim, pp. 649-667.
209. Becker, F.S., Schmoldt, H.-U., Adams, T.M., Wilhelm, S., and Kolmar, H. (2004) Ultrahigh-throughput screening based on cell-surface display and fluorescence-activated cell sorting or the identification of novel biocatalysts. Curr. Opin. Biotechnol., 15, 323-329;
210. Olsen, M.J., Gam, J., Iverson, B.L., and Georgiou, G., High-throughput FACS method for directed evolution of substrate specificity, in Methods in Molecular Biology, vol. 230 (eds F.H. Arnold and G. Georgiou), Humana Press Inc., Totowa, NJ, pp. 329-342.
211. Dröge, M.J., Boersma, Y.L., Van Pouderoyen, G., Vrenken, T.E., Rüggeberg, C.J., Reetz, M.T., Dijkstra, B.W., and Quax, W.J. (2006) Directed evolution of Bacillus subtilis lipase a by use of enantiomeric phosphonate inhibitors: crystal structures and phage display selection. ChemBioChem, 7, 149-157.
212. Lipovšek, D., Antipov, E., Armstrong, K.A., Olsen, M.J., Klibanov, A.M., Tidor, B., and Wittrup, K.D. (2007) Selection of horseradish peroxidase variants with enhanced enantioselectivity by yeast surface display. Chem. Biol., 14, 1176-1185.
213. Becker, S., Höbenreich, H., Vogel, A., Knorr, J., Wilhelm, S., Rosenau, F., Jaeger, K.-E., Reetz, M.T., and Kolmar, H. (2008) Single-cell high-throughput screening to identify enantioselective hydrolytic enzymes. Angew. Chem., 120, 5163-5166; Angew. Chem. Int. Ed. Engl., 47, 5085-5088.
214. Liese, A., Seelbach, K., and Wandrey, C. (2006) Industrial Biotransformations, Wiley-VCH Verlag GmbH, Weinheim.
215. Renugopalakrishnan, V., Garduño-Juárez, R., Narasimhan, G., Verma, C.S., Wei, X., and Li, P. (2005) Rational design of thermally stable proteins: relevance to bionanotechnology. J. Nanosci. Nanotechnol., 5, 1759-1767.
216. Gatti-Lafranconi, P., Caldarazzo, S.M., Villa, A., Alberghina, L., and Lotti, M. (2008) Unscrambling thermal stability and temperature adaptation in evolved variants of a cold-active lipase. FEBS Lett., 582, 2313-2318;
217. Frieden, C. (2007) Protein aggregation processes: in search of the mechanism. Protein Sci., 16, 2334-2344;
218. Ahmad, S. and Rao, N.M. (2009) Thermally denatured state determines refolding in lipase:mutational analysis. Protein Sci., 18, 1183-1196.
219. Martin, A., Schmid, F.X., and Sieber, V. (2003) PROSIDE-a phage-based method for selecting thermostable proteins, in Directed Enzyme Evolution: Screening and Selection Methods, vol. 230 (eds F.H. Arnold and G. Georgiou), Humana Press, Totowa, NJ, pp. 57-70.
220. Hecky, J., Mason, J.M., Arndt, K.M., and Müller, K.M. (2007) A general method of terminal truncation, evolution, and reelongation to generate enzymes of enhanced stability, in Protein Engineering Protocols, vol. 352 (eds K.M. Arndt and K.M.Müller), Humana Press, Totowa, NJ, pp. 275-304.
221. Valinger, G., Hermann, M., Wagner, U.G., and Schwab, H. (2007) Stability and activity improvement of cephalosporin esterase EstB from Burkholderia gladioli by directed evolution and structural interpretation of muteins. J. Biotechnol., 129, 98-108.
222. Petersen, E.I., Valinger, G., Sölkner, B., Stubenrauch, G., and Schwab, H. (2001) A novel esterase from Burkholderia gladioli which shows high deacetylation activity on cephalosporins is related to b-lactamases and DD-peptidases. J. Biotechnol., 89, 11-25.
223. Asako, H., Shimizu, M., and Itoh, N. (2008) Engineering of NADPHdependent aldo-keto reductase from Penicillium citrinum by directed evolution to improve thermostability and enantioselectivity. Appl. Microbiol. Biotechnol., 80, 805-812.
224. Palackal, N., Brennan, Y., Callen, W.N., Dupree, P., Frey, G., Goubet, F., Hazlewood, G.P., Healey, S., Kang, Y.E., Kretz, K.A., Lee, E., Tan, X., Tomlinson, G.L., Verruto, J., Wong, V.W.K., Mathur, E.J., Short, J.M., Robertson, D.E., and Steer, B.A. (2004) An evolutionary route to xylanase process fitness. Protein Sci., 13, 494-503;
225. Ruller, R., Deliberto, L., Lopes Ferreira, T., and Ward, R.J. (2007) Thermostable variants of the recombinant xylanase A from Bacillus subtilis produced by directed evolution show reduced heat capacity changes. Proteins, 70, 1280-1293.
226. Dumon, C., Varvak, A., Wall, M.A., Flint, J.E., Lewis, R.J., Lakey, J.H., Morland, C., Luginbühl, P., Healey, S., Todaro, T., Desantis, G., Sun, M., Parra-Gessert, L., Tan, X., Weiner, D.P., and Gilbert, H.J. (2008) Engineering hyperthermostability into a GH11 xylanase is mediated by subtle changes to protein structure. J. Biol. Chem., 283, 22557-22564;
227. Funke, S.A., Eipper, A., Reetz, M.T., Otte, N., Thiel, W., Van Pouderoyen, G., Dijkstra, B.W., Jaeger, K.-E., and Eggert, T. (2003) Directed evolution of an enantioselective Bacillus subtilis lipase. Biocatal. Biotransform., 21, 67-73.
228. Tang, S.-Y., Le, Q.-T., Shim, J.-H., Yang, S.-J., Auh, J.-H., Park, C., and Park, K.-H. (2006) Enhancing thermostability of maltogenic amylase from Bacillus thermoalkalophilus ET2 by DNA shuffling. FEBS J., 273, 3335-3345.
229. Kotzia, G.A. and Labrou, N.E. (2009) Engineering thermal stability of Lasparaginase by in vitro directed evolution. FEBS J., 276, 1750-1761.
230. Reetz, M.T., Soni, P., Acevedo, J.P., and Sanchis, J. (2009) Creation of an amino acid network of structurally coupled residues in the directed evolution of a thermostable enzyme. Angew. Chem., 121, 8418-8422; Angew. Chem. Int. Ed. Engl., 48, 8268-8272.
231. Reetz, M.T., Soni, P., and Fernández, L. (2009) Knowledge-guided laboratory evolution of protein thermolability. Biotechnol. Bioeng., 102, 1712-1717.
232. Mclachlan, M.J., Johannes, T.W., and Zhao, H. (2008) Further improvement of phosphite dehydrogenase thermostability by saturation mutagenesis. Biotechnol. Bioeng., 99, 268-274.
233. Heinzelman, P., Snow, C.D., Wu, I., Nguyen, C., Villalobos, A., Govindarajan, S., Minshull, J., and Arnold, F.H. (2009) A family of thermostable fungal cellulases created by structure-guided recombination. Proc. Natl. Acad. Sci. U.S.A., 106, 5610-5615.
234. Minagawa, H., Yoshida, Y., Kenmochi, N., Furuichi, M., Shimada, J., and Kaneko, H. (2007) Improving the thermal stability of lactate oxidase by directed evolution. Cell. Mol. Life Sci., 64, 77-81.
235. Stevenson, B.J., Liu, J.-W., and Ollis, D.L. (2008) Directed evolution of yeast pyruvate decarboxylase 1 for attenuated regulation and increased stability. Biochemistry, 47, 3013-3025.
236. Klibanov, A.M. (2001) Improving enzymes by using them in organic solvents. Nature, 409, 241-246;
237. Carrea, G. and Riva, S. (2008) Organic Synthesis with Enzymes in Non-aqueous Media, Wiley-VCH Verlag GmbH, Weinheim.
238. Zumárraga, M., Bulter, T., Shleev, S., Polaina, J., Martínez-Arias, A., Plou, F.J., Ballesteros, A., and Alcalde, M. (2007) In vitro evolution of a fungal laccase in high concentrations of organic cosolvents. Chem. Biol., 14, 1052-1064.
239. You, C., Huang, Q., Xue, H., Xu, Y., and Lu, H. (2009) Potential hydrophobic interaction between two cysteines in interior hydrophobic region improves thermostability of a family 11 xylanase from Neocallimastix patriciarum. Biotechnol. Bioeng., 105, 861-870.
240. Francotte, E. and Lindner, W. (2006) Chirality in Drug Research, Wiley-VCH Verlag GmbH, Weinheim.
241. Martín-Matute, B. and Bäckvall, J.-E. (2007) Dynamic kinetic resolution catalyzed by enzymes and metals. Curr. Opin. Chem. Biol., 11, 226-232.
242. Carballeira, J.D., Krumlinde, P., Bocola, M., Vogel, A., Reetz, M.T., and Bäckvall, J.-E. (2007) Directed evolution and axial chirality: optimization of the enantioselectivity of Pseudomonas aeruginosa lipase towards the kinetic resolution of a racemic allene. Chem. Commun., 1913-1915.
243. Suen, W.-C., Zhang, N., Xiao, L., Madison, V., and Zaks, A. (2004) Improved activity and thermostability of Candida antarctica lipase B by DNA family shuffling. Protein Eng., Design & Select., 17, 133-140.
244. Schmidt, M., Hasenpusch, D., Kähler, M., Kirchner, U., Wiggenhorn, K., Langel, W., and Bornscheuer, U.T. (2006) Directed evolution of an esterase fromPseudomonas fluorescens yields a mutant with excellent enantioselectivity and activity for the kinetic resolution of a chiral building block. ChemBioChem, 7, 805-809.
245. Bartsch, S., Kourist, R. and Bornscheuer, U.T. (2008) Complete inversion of enantioselectivity towards acetylated tertiary alcohols by a double mutant of a Bacillus subtilis esterase. Angew. Chem., 120, 1531-1534; Angew. Chem. Int. Ed. Engl., 47, 1508-1511.
246. Kotik, M., štěpánek, V., Kyslík, P., and Marešová, H. (2007) Cloning of an epoxide hydrolase-encoding gene from Aspergillus niger M200, overexpression in E. coli, and modification of activity and enantioselectivity of the enzyme by protein engineering. J. Biotechnol., 132, 8-15.
247. Van Loo, B., Spelberg, J.H.L., Kingma, J., Sonke, T., Wubbolts, M.G., and Janssen, D.B. (2004) Directed evolution of epoxide hydrolase from A. radiobacter toward higher enantioselectivity by error-pronePCRand DNA shuffling. Chem. Biol., 11, 981-990.
248. Kotik, M., Archelas, A., Famerová, V., Qubrechtová, P., and Kren V., (2011) Laboratory evolution of an epoxide hydrolase - towards an enantioconvergent biocatalyst J. Biotechnol., 156, 1-10.
249. DeSantis, G., Wong, K., Farwell, B., Chatman, K., Zhu, Z., Tomlinson, G., Huang, H., Tan, X., Bibbs, L., Chen, P., Kretz, K., and Burk, M.J. (2003) Creation of a productive, highly enantioselective nitrilase through gene site saturation mutagenesis (GSSM). J. Am. Chem. Soc., 125, 11476-11477.
250. Hill, C.M., Li, W.-S., Thoden, J.B., Holden, H.M., and Raushel, F.M. (2003) Enhanced degradation of chemical warfare agents through molecular engineering of the phosphotriesterase active site. J. Am. Chem. Soc., 125, 8990-8991.
251. Fong, S., Machajewski, T.D., Mak, C.C., and Wong, C.-H. (2000) Directed evolution of D-2-keto-3-deoxy-6-phosphogluconate aldolase to new variants for the efficient synthesis of D-and L-Sugars. Chem. Biol., 7, 873-883.
252. Wada, M., Hsu, C.-C., Franke, D., Mitchell, M., Heine, A., Wilson, I., and Wong, C.-H. (2003) Directed evolution of N-acetylneuraminic acid aldolase to catalyze enantiomeric aldol reactions. Bioorg. Med. Chem., 11, 2091-2098.
253. Williams, G.J., Woodhall, T., Farnsworth, L.M., Nelson, A., and Berrry, A. (2006) Creation of a pair of stereochemically complementary biocatalysts. J. Am. Chem. Soc., 128, 16238-16247.
254. Williams, G.J., Domann, S., Nelson, A., and Berry, A. (2003) Modifying the stereochemistry of an enzyme-catalyzed reaction by directed evolution. Proc. Natl. Acad. Sci. U.S.A., 100, 3143-3148.
255. Jennewein, S., Schürmann, M., Wolberg, M., Hilker, I., Luiten, R., Wubbolts, M., and Mink, D. (2006) Directed evolution of an industrial biocatalyst: 2-deoxy-D-ribose 5-phosphate aldolase. Biotechnol. J., 1, 537-548.
256. Ran, N., Draths, K.M., and Frost, J.W. (2004) Creation of a shikimate pathway variant. J. Am. Chem. Soc., 126, 6856-6857.
257. Ran, N. and Frost, J.W. (2007) Directed evolution of 2-keto-3-deoxy-6-phosphogalactonate aldolase to replace 3-deoxy-D-arabino-heptulosonic acid 7-phosphate synthase. J. Am. Chem. Soc., 129, 6130-6139.
258. Lingen, B., Kolter-Jung, D., Dünkelmann, P., Feldmann, R., Grötzinger, J., Pohl, M., and Müller, M. (2003) Alteration of the substrate specificity of benzoylformate decarboxylase from Pseudomonas putida by directed evolution. ChemBioChem, 4, 721-726.
259. Liang, J., Lalonde, J., Borup, B., Mitchell, V., Mundorff, E., Trinh, N., Kochrekar, D.A., Cherat, R.N. and Pai, G.G. (2010) Development of biocatalytic process as an alternative to the ()-DIP-Cl-mediated asymmetric reduction of a key intermediate of montelukast. Org. Process Res. Dev., 14, 193-198;
260. Liang, J., Mundorff, E., Voladri, R., Jenne, S., Gilson, L., Conway, A., Krebber, A., Wong, J., Huismann, G., Truesdale, S., and Lalonde, J. (2010)Highly enantioselective reduction of a small heterocyclic ketone: biocatalytic reduction of tetrahydrothiophene-3-one to the corresponding (R)-alcohol. Org. Process Res. Dev., 14, 188-192.
261. Reetz, M.T., Brunner, B., Schneider, T., Schulz, F., Clouthier, C.M., and Kayser, M.M. (2004) Directed evolution as a method to create enantioselective cyclohexanone monooxygenases for catalysis in Baeyer-Villiger reactions. Angew. Chem., 116, 4167-4170; Angew. Chem. Int. Ed., 43, 4075-4078;
262. Mihovilovic, M.D., Rudroff, F., Winninger, A., Schneider, T., Schulz, F., and Reetz, M.T. (2006) Microbial Baeyer-Villiger oxidation: stereopreference and substrate acceptance of cyclohexanone monooxygenase mutants prepared by directed evolution. Org. Lett., 8, 1221-1224.
263. Reetz, M.T. and Wu, S. (2009) Laboratory evolution of robust and enantioselective Baeyer-Villiger monooxygenases for asymmetric catalysis. J. Am. Chem. Soc., 131, 15424-15432.
264. Wu, S., Acevedo, J.P., and Reetz, M.T. (2010) Induced allostery in the directed evolution of an enantioselective Baeyer-Villiger monooxygenase. Proc. Natl. Acad. Sci. U.S.A., 107, 2775-2780.
265. Kirschner, A. and Bornscheuer, U.T. (2008) Directed evolution of a Baeyer-Villiger monooxygenase to enhance enantioselectivity. Appl. Microbiol. Biotechnol., 81, 465-472.
266. Peters, M.W., Meinhold, P., Glieder, A., and Arnold, F.H. (2003) Regio-and enantioselective alkane hydroxylation with engineered cytochromes P450 BM-3. J. Am. Chem. Soc., 125, 13442-13450;
267. Landwehr, M., Hochrein, L., Otey, C.R., Kasrayan, A., Bäckvall, J.-E., and Arnold, F.H. (2006) Enantioselective a-hydroxylation of 2-arylacetic acid derivatives and Buspirone catalyzed by engineering cytochrome P450 BM-3. J. Am. Chem. Soc., 128, 6058-6059;
268. Urlacher, V.B., Lutz-Wahl, S. and Schmid, R.D. (2004) Microbial P450 enzymes in biotechnology. Appl. Microbiol. Biotechnol., 64, 317-325;
269. Gillam, E.M.J. (2008) Engineering cytochrome P450 enzymes. Chem. Res. Toxicol., 21, 220-231;
270. Münzer, D.F., Meinhold, P., Peters, M.W., Feichtenhofer, S., Griengl, H., Arnold, F.H., Glieder, A., and De Raadt, A. (2005) Stereoselective hydroxylation of an achiral cyclopentanecarboxylic acid derivative using engineered P450s BM-3. Chem. Commun., 2597-2599;
271. Kille, S., Zilly, F.E., Acevedo, J.P., and Reetz, M.T. (2011) Regio-and stereoselectivity of P450-catalysed hydroxylation of steroids controlled by laboratory evolution. Nat. Chem., 3, 738-743.
272. Antipov, E., Cho, A.E., Wittrup, K.D., and Klibanov, A.M. (2008) Highly L and D enantioselective variants of horseradish peroxidase discovered by an ultrahighthroughput selection method. Proc. Natl. Acad. Sci. U.S.A., 105, 17694-17699.
273. Alexeeva, M., Enright, A., Dawson, M.J., Mahmoudian, M. and Turner, N.J. (2002) Deracemization of a-methylbenzylamine using an enzyme obtained by in vitro evolution. Angew. Chem., 114, 3309-3312; Angew. Chem. Int. Ed., 41, 3177-3180;
274. Carr, R., Alexeeva, M., Enright, A., Eve, T.S.C., Dawson, M.J., and Turner, N.J. (2003) Directed evolution of an amine oxidase possessing both broad substrate specificity and high enantioselectivity. Angew. Chem., 115, 4955-4958; Angew. Chem. Int. Ed., 42, 4807-4810.
275. Avi, M., Wiedner, R.M., Griengl, H., and Schwab, H. (2008) Improvement of a stereoselective biocatalytic synthesis by substrate and enzyme engineering: 2-hydroxy-(40-oxocyclohexyl)acetonitrile as the model. Chem. Eur. J., 14, 11415-11422.
276. Jacobsen, E.N. (2000) Asymmetric catalysis of epoxide ring-opening reactions. Acc. Chem. Res., 33, 421-431.
277. Zou, J.Y., Hallberg, B.M., Bergfors, T., Oesch, F., Arand, M., Mowbray, S.L., and Jones, T.A. (2000) Structure of Aspergillus niger epoxide hydrolase at 1.8 Aresolution: implications for the structure and function of the mammalian microsomal class of epoxide hydrolases. Structure (London), 8, 111-122.
278. Reetz, M.T., Bocola, M., Wang, L.-W., Sanchis, J., Cronin, A., Arand, M., Zou, J., Archelas, A., Bottalla, A.-L., Naworyta, A., and Mowbray, S.L. (2009) Directed evolution of an enantioselective epoxide hydrolase: uncovering the source of enantioselectivity at each evolutionary stage. J. Am. Chem. Soc., 131, 7334-7343.
279. Kayser, M.M. (2009) Designer reagents recombinant microorganisms: new and powerful tools for organic synthesis. Tetrahedron, 65, 947-974;
280. Mihovilovic, M.D. (2006) Enzyme mediated Baeyer-Villiger oxidations. Curr. Org. Chem., 10, 1265-1287.
281. Bocola, M., Schulz, F., Leca, F., Vogel, A., Fraaije, M.W., and Reetz, M.T. (2005) Converting phenylacetone monooxygenase into phenylcyclohexanone monooxygenase by rational design: towards practical Baeyer-Villiger monooxygenases. Adv. Synth. Catal., 347, 979-986.
282. Fraaije, M.W., Wu, J., Heuts, D.P.H.M., Van Hellemond, E.W., Spelberg, J.H.L., and Janssen, D.B. (2005) Discovery of a thermostable Baeyer-Villiger monooxygenase by genome mining. Appl. Microbiol. Biotechnol., 66, 393-400;
283. Zambianchi, F., Fraaije, M.W., Carrea, G., De Gonzalo, G., Rodríguez, C., Gotor, V., and Ottolina, G. (2007) Titration and assignment of residues that regulate the enantioselectivity of phenylacetone monooxygenase. Adv. Synth. Catal., 349, 1327-1331;
284. Rodrígues, C., DeGonzalo, G., Torres Pazmiño, D.E., Fraaije, M.W., and Gotor, V. (2008) Selective Baeyer- Villiger oxidation of racemic ketones in aqueous-organic media catalyzed by phenylacetone monooxygenase. Tetrahedron: Asymmetry, 19, 197-203.
285. Malito, E., Alfieri, A., Fraaije, M.W., and Mattevi, A. (2004) Crystal structure of a Baeyer-Villiger monooxygenase. Proc. Natl. Acad. Sci. U.S.A., 101, 13157-13162.
286. Koeller, K.M. and Wong, C.-H. (2001) Enzymes for chemical synthesis. Nature, 409, 232-240.
287. Bolt, A., Berry, A., and Nelson, A. (2008) Directed evolution of aldolases for exploitation in synthetic organic chemistry. Arch. Biochem. Biophys., 474, 318-330.
288. Tokuriki, N. and Tawfik, D.S. (2009) Chaperonin overexpression promotes genetic variation and enzyme evolution. Nature, 459, 668-673;
289. Matsumura, I. and Ivanov, A.A. (2009) How evolving enzymes can beat the heat and avoid defeat. Nat. Chem. Biol., 5, 538-539.
290. Glieder, A., Weis, R., Skranc, W., Poechlauer, P., Dreveny, I., Majer, S., Wubbolts, M., Schwab, H., andGruber, K. (2003) Comprehensive step-by-step engineering of an (R)-hydroxynitrile lyase for large-scale asymmetric synthesis. Angew. Chem., 115, 4963-4966; Angew. Chem. Int. Ed., 42, 4815-4818.
291. Khersonsky, O., Röthlisberger, D., Wollacott, A.M., Murphy, P., Dym, O., Albeck, S., Kiss, G., Houk, K.N., Baker, D., and Tawfik, D.S. (2011) Optimization of the in-silico-designed Kemp eliminase KE70 by computational design and directed evolution. J. Mol. Biol., 407, 391-412.