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Modular enzymes

Abstract

Although modular macromolecular devices are encountered frequently in a variety of biological situations, their occurrence in biocatalysis has not been widely appreciated. Three general classes of modular biocatalysts can be identified: enzymes in which catalysis and substrate specificity are separable, multisubstrate enzymes in which binding sites for individual substrates are modular, and multienzyme systems that can catalyse programmable metabolic pathways. In the postgenomic era, the discovery of such systems can be expected to have a significant impact on the role of enzymes in synthetic and process chemistry.

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Figure 1: Separate catalytic and molecular recognition domains of the Fok I restriction endonuclease.
Figure 2: Recognition by penicillin G acylase of a modular chemical feature (the phenylacetyl group) in an otherwise generic substrate.
Figure 3: Separation of molecular-recognition features in modular multisubstrate enzymes.
Figure 4: Separation of electrophile and nucleophile recognition in modules of polyketide synthases and non-ribosomal peptide synthetases.
Figure 5: Introduction of auxiliary catalytic domains into the module of a polyketide synthase.
Figure 6: Modular protein–protein interactions in the selective channelling of intermediates between successive catalysts in a multistep metabolic pathway.

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Elizabeth L. Bell, William Finnigan, … Sabine L. Flitsch

References

  1. Crick, F. H., Barnett, I., Brenner, S. & Watts-Tobin, R. General nature of the genetic code for proteins. Nature 192, 1227–1232 (1961).

    Article  ADS  CAS  Google Scholar 

  2. Cohen, G. B., Sen, R. & Baltimore, D. Modular binding proteins in signal transduction proteins . Cell 80, 237–248 (1995).

    Article  CAS  Google Scholar 

  3. Sudol, M. From Src homology domains to other signaling modules: proposal of the protein recognition code. Oncogene 17, 1469– 1474 (1998).

    Article  CAS  Google Scholar 

  4. Frankel, A. D. & Kim, P. S. Modular structure of transcription factors: implications for gene regulation. Cell 65, 717–719 (1991).

    Article  CAS  Google Scholar 

  5. Ptashne, M. & Gann, A. Imposing specificity by localization: mechanism and evolvability. Curr. Biol. 8, R897–R904 (1998).

    Article  CAS  Google Scholar 

  6. Rashin, A. A. Location of domains in globular proteins. Nature 291 , 85–87 (1981).

    Article  ADS  CAS  Google Scholar 

  7. Orengo, C. A., Jones, D. T. & Thornton, J. M. Protein superfamilies and domain superfolds. Nature 372, 631–634 ( 1994).

    Article  ADS  CAS  Google Scholar 

  8. Henikoff, S. et al. Gene families: the taxonomy of protein paralogs and chimeras . Science 278, 609–614 (1997).

    Article  ADS  CAS  Google Scholar 

  9. Riley, M. & Labedan, B. Protein evolution viewed through Escherichia coli protein sequences: introducing the notion of a structural segment of homology, the module. J. Mol. Biol. 268, 857–868 (1997).

    Article  CAS  Google Scholar 

  10. Dalal, S., Balasubramanian, S. & Regan, L. Protein alchemy: changing β-sheet into α-helix . Nature Struct. Biol. 4, 548– 552 (1997).

    Article  CAS  Google Scholar 

  11. Jencks, W. P. Catalysis in Chemistry and Enzymology 615–806 (Dover, New York, 1987).

    Google Scholar 

  12. Li, L., Wu, L. P. & Chandrasegaran, S. Functional domains in Fok-I restriction endonuclease . Proc. Natl Acad. Sci. USA 89, 4275– 4279 (1992).

    Article  ADS  CAS  Google Scholar 

  13. Kim, Y. G., Cha, J. & Chandrasegaran, S. Hybrid restriction enzymes—zinc finger fusions to Fok I cleavage domain. Proc. Natl Acad. Sci. USA 93, 1156–1160 (1996).

    Article  ADS  CAS  Google Scholar 

  14. Huang, B. H., Schaeffer, C. J., Li, Q. H. & Tsai, M. D. Splase—a new class I zinc-finger restriction endonuclease with specificity for Sp1 binding sites. J. Protein Chem. 15, 481–489 (1996).

    Article  CAS  Google Scholar 

  15. Aggarwal, A. K. & Wah, D. A. Novel site-specific DNA endonucleases. Curr. Opin. Struct. Biol. 8, 19–25 (1998).

    Article  CAS  Google Scholar 

  16. Fernandes, A. C. et al. Homologous xylanases from Clostridium thermocellum: evidence for bi-functional activity, synergism between xylanase catalytic modules and the presence of xylan-binding domains in enzyme complexes. Biochem. J. 342, 105–110 (1999).

    CAS  PubMed  PubMed Central  Google Scholar 

  17. Bolam, D. N. et al. Pseudomonas cellulose-binding domains mediate their effects by increasing enzyme substrate proximity. Biochem. J. 331, 775–781 (1998).

    Article  CAS  Google Scholar 

  18. Marcotte, E. M. et al. Detecting protein function and protein-protein interactions from genome sequences. Science 285, 751– 753 (1999).

    Article  CAS  Google Scholar 

  19. Jencks, W. P. On the attribution and additivity of binding energies. Proc. Natl Acad. Sci. USA 78, 4046–4050 (1981).

    Article  ADS  CAS  Google Scholar 

  20. Murphy, F. L. & Cech, T. R. Alteration of substrate specificity for the endoribonucleolytic cleavage of RNA by the Tetrahymena ribozyme . Proc. Natl Acad. Sci. USA 86, 9218– 9222 (1989).

    Article  ADS  CAS  Google Scholar 

  21. Liu, Y., Shah, K., Yang, F., Witucki, L. & Shokat, K. M. Engineering Src family protein kinases with unnatural nucleotide specificity. Chem. Biol. 5, 91 –101 (1998).

    Article  CAS  Google Scholar 

  22. Hedstrom, L., Farrjones, S., Kettner, C. A. & Rutter, W. J. Converting trypsin to chymotrypsin—ground-state binding does not determine substrate specificity. Biochemistry 33, 8764–8769 (1994).

    Article  CAS  Google Scholar 

  23. Stemmer, W. P. C. Rapid evolution of a protein in vitro by DNA shuffling. Nature 370, 389–391 ( 1994).

    Article  ADS  CAS  Google Scholar 

  24. Jaeger, K. E., Dijkstra, B. W. & Reetz, M. T. Bacterial biocatalysts: molecular biology, three-dimensional structures, and biotechnological applications of lipases. Annu. Rev. Microbiol. 53, 327–351 (1999).

    Article  Google Scholar 

  25. Pathak, T. & Waldmann, H. Enzymes and protecting group chemistry . Curr. Opin. Chem. Biol. 2, 112– 120 (1998).

    Article  CAS  Google Scholar 

  26. Costello, C. A., Kreuzman, A. J. & Zmijewski, M. J. Selective deprotection of phthalyl protected amines . Tetrahedron Lett. 37, 7469– 7472 (1996).

    Article  CAS  Google Scholar 

  27. Villarreal, D. T., Turco, R. F. & Konopka, A. A structure-activity study with aryl acylamidases. Appl. Env. Microbiol. 60, 3939–3944 (1994).

    CAS  Google Scholar 

  28. Roberts, S. M. Preparative biotransformations: the employment of enzymes and whole-cells in synthetic organic chemistry. J. Chem. Soc. Perkin Trans. 1, 157–169 (1998).

    Article  Google Scholar 

  29. Roberts, S. M. Preparative biotransformations. J. Chem. Soc. Perkin Trans. 1, 1–21 (1999).

    Article  Google Scholar 

  30. Roberts, S. M. Preparative biotransformations. J. Chem. Soc. Perkin Trans. 1, 611–633 (2000).

    Article  Google Scholar 

  31. Duggleby, H. J. et al. Penicillin acylase has a single amino-acid catalytic centre . Nature 373, 264–268 (1995).

    Article  ADS  CAS  Google Scholar 

  32. Done, S. H., Brannigan, J. A., Moody, P. C. E. & Hubbard, R. E. Ligand-induced conformational change in penicillin acylase. J. Mol. Biol. 284, 463–475 (1998).

    Article  CAS  Google Scholar 

  33. Lang, D. A., Mannesse, M. L. M., DeHaas, G. H., Verheij, H. M. & Dijkstra, B. W. Structural basis of the chiral selectivity of Pseudomonas cepacia lipase. Eur. J. Biochem. 254, 333–340 ( 1998).

    Article  CAS  Google Scholar 

  34. Fersht, A. Enzyme Structure and Mechanism (Freeman, 1985).

    Google Scholar 

  35. Goulding, C. W., Postigo, D. & Matthews, R. G. Cobalamin-dependent methionine synthase is a modular protein with distinct regions for binding homocysteine, methyltetrahydrofolate, cobalamin, and adenosylmethionine. Biochemistry 36, 8082–8091 (1997).

    Article  CAS  Google Scholar 

  36. Ha, S., Walker, D., Shi, Y. & Walker, S. The 1.9 A crystal structure of Escherichia coli MurG, a membrane-associated glycosyltransferase involved in peptidoglycan biosynthesis. Protein Sci. 9, 1045–1052 (2000).

    Article  CAS  Google Scholar 

  37. Campbell, R. E., Mosimann, S. C., van de Rijn, I., Tanner, M. E. & Strynadka, N. C. The first structure of UDP-glucose dehydrogenase reveals the catalytic residues necessary for the two-fold oxidation . Biochemistry 39, 7012– 7023 (2000).

    Article  CAS  Google Scholar 

  38. Borisova, S. A., Zhao, L., Sherman, D. H. & Liu, H. W. Biosynthesis of desosamine: construction of a new macrolide carrying a genetically designed sugar moiety. Org. Lett. 1, 133– 136 (1999).

    Article  CAS  Google Scholar 

  39. Olano, C., Lomovskaya, N., Fonstein, L., Roll, J. T. & Hutchinson, C. R. A two-plasmid system for the glycosylation of polyketide antibiotics: bioconversion of epsilon-rhodomycinone to rhodomycin D. Chem. Biol. 6, 845– 855 (1999).

    Article  CAS  Google Scholar 

  40. Kapitonov, D. & Yu, R. K. Conserved domains of glycosyltransferases . Glycobiology 9, 961–978 (1999).

    Article  CAS  Google Scholar 

  41. Cane, D. E., Walsh, C. T. & Khosla, C. Harnessing the biosynthetic code. Combinations, permutations, mutations. Science 282, 63– 68 (1998).

    Article  CAS  Google Scholar 

  42. Khosla, C., Gokhale, R., Jacobsen, J. R. & Cane, D. E. Tolerance and specificity of polyketide synthases. Annu. Rev. Biochem. 68, 219–253 ( 1999).

    Article  CAS  Google Scholar 

  43. Mootz, H. D. & Marahiel, M. A. Design and application of multimodular peptide synthetases. Curr. Opin. Biotechnol. 10, 341–348 (1999).

    Article  CAS  Google Scholar 

  44. Belshaw, P. J., Walsh, C. T. & Stachelhaus, T. Aminoacyl-CoAs as probes of condensation domain selectivity in nonribosomal peptide synthesis. Science 284, 486–489 (1999).

    Article  ADS  CAS  Google Scholar 

  45. Wu, N., Kudo, F., Cane, D. E. & Khosla, C. Analysis of the molecular recognition features of individual modules derived from the erythromycin polyketide synthase. J. Am. Chem. Soc. 122, 4847–4852 (2000).

    Article  CAS  Google Scholar 

  46. McDaniel, R. et al. Multiple genetic modifications of the erythromycin polyketide synthase to produce a library of novel “unnatural” natural products . Proc. Natl Acad. Sci. USA 96, 1846– 1851 (1999).

    Article  ADS  CAS  Google Scholar 

  47. Geck, M. K. & Kirsch, J. F. A novel, definitive test for substrate channeling illustrated with the aspartate aminotransferase/malate dehydrogenase system. Biochemistry 38, 8032– 8037 (1999).

    Article  CAS  Google Scholar 

  48. Hyde, C. C., Ahmed, S. A., Padlan, E. A., Miles, E. W. & Davies, D. R. Three-dimensional structure of the tryptophan synthase alpha 2 beta 2 multienzyme complex from Salmonella typhimurium. J. Biol. Chem. 263, 17857 –17871 (1988).

    CAS  PubMed  Google Scholar 

  49. Krahn, J. M. et al. Coupled formation of an amidotransferase interdomain ammonia channel and a phosphoribosyltransferase active site. Biochemistry 36, 11061–11068 ( 1997).

    Article  CAS  Google Scholar 

  50. Thoden, J. B., Holden, H. M., Wesenberg, G., Raushel, F. M. & Rayment, I. Structure of carbamoyl phosphate synthetase: a journey of 96 Å from substrate to product. Biochemistry 36, 6305–6316 (1997).

    Article  CAS  Google Scholar 

  51. Perham, R. N. & Reche, P. A. Swinging arms in multifunctional enzymes and the specificity of post-translational modification. Biochem. Soc. Trans. 26, 299–303 (1998).

    Article  CAS  Google Scholar 

  52. O'Shea, E. K., Rutkowski, R., Stafford, W. F. III & Kim, P. S. Preferential heterodimer formation by isolated leucine zippers from Fos and Jun. Science 245, 646 –648 (1989).

    Article  ADS  CAS  Google Scholar 

  53. Kraulis, P. J. Molscript—a program to produce both detailed and schematic plots of protein structures. J. Appl. Crystallogr. 24, 946–950 (1991).

    Article  Google Scholar 

  54. Kao, C. M. et al. Gain of function mutagenesis of a modular polyketide synthase II. Engineered biosynthesis of an eight-membered ring tetraketide lactone . J. Am. Chem. Soc. 119, 11339– 11340 (1997).

    Article  CAS  Google Scholar 

  55. Cortes, J., Haydock, S. F., Roberts, G. A., Bevitt, D. J. & Leadlay, P. F. An unusually large multifunctional polypeptide in the erythromycin-producing polyketide synthase of Saccharopolyspora erythraea. Nature 348, 176– 178 (1990).

    Article  ADS  CAS  Google Scholar 

  56. Donadio, S., Staver, M. J., McAlpine, J. B., Swanson, S. J. & Katz, L. Modular organization of genes required for complex polyketide biosynthesis. Science 252, 675–679 (1991).

    Article  ADS  CAS  Google Scholar 

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Acknowledgements

Research on modular enzymes in C.K.'s laboratory is supported by grants from the National Science Foundation and the National Institutes of Health. P.B.H. is a Terman Fellow, a Searle Scholar and a Burroughs–Wellcome Young Investigator in the Pharmacological Sciences. We thank S. Walker for helpful discussions regarding glycosyltransferases.

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Correspondence to Chaitan Khosla.

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Khosla, C., Harbury, P. Modular enzymes. Nature 409, 247–252 (2001). https://doi.org/10.1038/35051723

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