Skip to main content

Production of Selenomethionyl Proteins in Prokaryotic and Eukaryotic Expression Systems

  • Protocol
Book cover Macromolecular Crystallography Protocols

Part of the book series: Methods in Molecular Biology ((MIMB,volume 363))

Abstract

The use of selenomethionine as a phasing tool was first reported in 1990. Engineering of selenomethionyl proteins for structure determination is now routine. In fact, selenium is by far the most commonly used anomalous scatterer for multiwavelength anomalous diffraction studies. The past few years have seen new developments, which demonstrated the feasibility of expressing selenomethionyl protein in eukaryotic systems. In this chapter, the different methods available for producing selenomethionine-labeled proteins in bacteria, as well as in yeast and mammalian cells will be presented, along with tips for purifying and crystallizing selenomethionyl proteins.

This is a preview of subscription content, log in via an institution to check access.

Access this chapter

Protocol
USD 49.95
Price excludes VAT (USA)
  • Available as PDF
  • Read on any device
  • Instant download
  • Own it forever
eBook
USD 84.99
Price excludes VAT (USA)
  • Available as EPUB and PDF
  • Read on any device
  • Instant download
  • Own it forever
Softcover Book
USD 109.00
Price excludes VAT (USA)
  • Compact, lightweight edition
  • Dispatched in 3 to 5 business days
  • Free shipping worldwide - see info
Hardcover Book
USD 109.99
Price excludes VAT (USA)
  • Durable hardcover edition
  • Dispatched in 3 to 5 business days
  • Free shipping worldwide - see info

Tax calculation will be finalised at checkout

Purchases are for personal use only

Institutional subscriptions

References

  1. Hendrickson, W. A., Horton, J. R. and LeMaster, D. M. (1990) Selenomethionyl proteins produced for analysis by multiwavelength anomalous diffraction (MAD): a vehicle for direct determination of three-dimensional structure. EMBO J. 9, 1665ā€“1672.

    CASĀ  PubMedĀ  Google ScholarĀ 

  2. Bernard, A. R., Wells, T. N., Cleasby, A., Borlat, F., Payton, M. A., and Proudfoot, A. E. (1995). Selenomethionine labelling of phosphomannose isomerase changes its kinetic properties. Eur. J. Biochem. 230, 111ā€“118.

    ArticleĀ  CASĀ  PubMedĀ  Google ScholarĀ 

  3. Gassner, N. C., Baase, W. A., Hausrath, A. C., and Matthews, B. W. (1999) Substitution with selenomethionine can enhance the stability of methionine-rich proteins. J. Mol. Biol. 294, 17ā€“20.

    ArticleĀ  CASĀ  PubMedĀ  Google ScholarĀ 

  4. Jiang, J. and Sweet, R. M. (2004) Protein Data Bank depositions from synchrotron sources. J. Synchrotron. Radiat. 11, 319ā€“327.

    ArticleĀ  CASĀ  PubMedĀ  Google ScholarĀ 

  5. Sambrook, J. and Russell, D. W. (2001) Molecular Cloning: A Laboratory Manual, Cold Spring Harbor Laboratory Press, Cold Spring Harbor, NY.

    Google ScholarĀ 

  6. Sreenath, H. K., Bingman, C. A., Buchan, B. W., et al. (2005) Protocols for production of selenomethionine-labeled proteins in 2-L polyethylene terephthalate bottles using auto-induction medium. Protein Expr. Purif. 40, 256ā€“267.

    ArticleĀ  CASĀ  PubMedĀ  Google ScholarĀ 

  7. Cowie, D. B. and Cohen, G. N. (1957) Biosynthesis by E. coli of active altered proteins containing selenium instead of sulfur. Biochim. Biophys. Acta 26, 252ā€“261.

    ArticleĀ  CASĀ  PubMedĀ  Google ScholarĀ 

  8. DoubliĆ©, S. (1997) Preparation of selenomethionyl proteins for phase determination. Methods Enzymol. 276, 523ā€“530.

    ArticleĀ  PubMedĀ  Google ScholarĀ 

  9. Berne, P. F., DoubliƩ, S., and Carter, C. W., Jr. (1999) Molecular biology for structural biology. In: Crystallization of Nucleic Acids and Proteins, 2nd ed., (Ducruix, A. and GiegƩ, R., eds.), Oxford University Press, Oxford, UK.

    Google ScholarĀ 

  10. Van Duyne, G. D., Standaert, R. F., Karplus, P. A., Schreiber, S. L. and Clardy, J. (1993) Atomic structures of the human immunophilin FKBP-12 complexes with FK506 and Rapamycin. J. Mol. Biol. 229, 105ā€“124.

    ArticleĀ  PubMedĀ  Google ScholarĀ 

  11. Studier, F. W. (2005) Protein production by auto-induction in high density shaking cultures. Protein Expr. Purif. 41, 207ā€“234.

    ArticleĀ  CASĀ  PubMedĀ  Google ScholarĀ 

  12. Bushnell, D. A., Cramer, P., and Kornberg, R. D. (2001) Selenomethionine incorporation in Saccharomyces cerevisiae RNA polymerase II. Structure (Camb) 9, R11ā€“R14.

    ArticleĀ  CASĀ  Google ScholarĀ 

  13. Larsson, A. M., Stahlberg, J., and Jones, T. A. (2002) Preparation and crystallization of selenomethionyl dextranase from Penicillium minioluteum expressed in Pichia pastoris. Acta Crystallogr. D Biol. Crystallogr. 58, 346ā€“348.

    ArticleĀ  PubMedĀ  Google ScholarĀ 

  14. Xu, B., Munoz, I. I., Janson, J. C., and Stahlberg, J. (2002) Crystallization and X-ray analysis of native and selenomethionyl beta-mannanase Man5A from blue mussel, Mytilus edulis, expressed in Pichia pastoris. Acta Crystallogr. D Biol. Crystallogr. 58, 542ā€“545.

    ArticleĀ  PubMedĀ  Google ScholarĀ 

  15. Larsson, A. M., Andersson, R., Stahlberg, J., Kenne, L., and Jones, T. A. (2003) Dextranase from Penicillium minioluteum: reaction course, crystal structure, and product complex. Structure (Camb) 11, 1111ā€“1121.

    ArticleĀ  CASĀ  Google ScholarĀ 

  16. Cramer, P., Bushnell, D. A., Fu, J., et al. (2000) Architecture of RNA polymerase II and implications for the transcription mechanism. Science 288, 640ā€“649.

    ArticleĀ  CASĀ  PubMedĀ  Google ScholarĀ 

  17. McWhirter, S. M., Pullen, S. S., Holton, J. M., Crute, J. J., Kehry, M. R., and Alber, T. (1999). Crystallographic analysis of CD40 recognition and signaling by human TRAF2. Proc. Natl. Acad. Sci. USA 96, 8408ā€“8413.

    ArticleĀ  CASĀ  PubMedĀ  Google ScholarĀ 

  18. Aertgeerts, K., Ye, S., Tennant, M. G., et al. (2004) Crystal structure of human dipeptidyl peptidase IV in complex with a decapeptide reveals details on substrate specificity and tetrahedral intermediate formation. Protein Sci, 13, 412ā€“421.

    ArticleĀ  CASĀ  PubMedĀ  Google ScholarĀ 

  19. Bellizzi, J. J., Widom, J., Kemp, C. W., and Clardy, J. (1999) Producing selenomethionine-labeled proteins with a baculovirus expression vector system. Structure Fold. Des. 7, R263ā€“R267.

    ArticleĀ  CASĀ  PubMedĀ  Google ScholarĀ 

  20. Ogiso, H., Ishitani, R., Nureki, O., et al. (2002) Crystal structure of the complex of human epidermal growth factor and receptor extracellular domains. Cell 110, 775ā€“787.

    ArticleĀ  CASĀ  PubMedĀ  Google ScholarĀ 

  21. Hamaoka, B. Y., Dann, C. E., 3rd, Geisbrecht, B. V., and Leahy, D. J. (2004) Crystal structure of Caenorhabditis elegans HER-1 and characterization of the interaction between HER-1 and TRA-2A. Proc. Natl. Acad. Sci. USA 101, 11,673ā€“11,678.

    ArticleĀ  CASĀ  PubMedĀ  Google ScholarĀ 

  22. Davis, S. J., Ikemizu, S., Collins, A. V., et al. (2001) Crystallization and functional analysis of a soluble deglycosylated form of the human costimulatory molecule B7-1. Acta Crystallogr. D Biol. Crystallogr. 57, 605ā€“608.

    ArticleĀ  CASĀ  PubMedĀ  Google ScholarĀ 

  23. Carfi, A., Smith, C. A., Smolak, P. J., McGrew, J., and Wiley, D. C. (1999) Structure of a soluble secreted chemokine inhibitor vCCI (p35) from cowpox virus. Proc. Natl. Acad. Sci. USA 96, 12,379ā€“12,383.

    ArticleĀ  CASĀ  PubMedĀ  Google ScholarĀ 

  24. Wu, H., Lustbader, J. W., Liu, Y., Canfield, R. E., and Hendrickson, W. A. (1994) Structure of human chorionic gonadotropin at 2.6 A resolution from MAD analysis of the selenomethionyl protein. Structure 2, 545ā€“558.

    ArticleĀ  CASĀ  PubMedĀ  Google ScholarĀ 

  25. Lustbader, J. W., Wu, H., Birken, S., et al. (1995) The expression, characterization, and crystallization of wild-type and selenomethionyl human chorionic gonadotropin. Endocrinology 136, 640ā€“650.

    ArticleĀ  CASĀ  PubMedĀ  Google ScholarĀ 

  26. May, A. P., Robinson, R. C., Aplin, R. T., Bradfield, P., Crocker, P. R., and Jones, E. Y. (1997) Expression, crystallization, and preliminary X-ray analysis of a sialic acid-binding fragment of sialoadhesin in the presence and absence of ligand. Protein Sci. 6, 717ā€“721.

    ArticleĀ  CASĀ  PubMedĀ  Google ScholarĀ 

  27. Wally, J., Halbrook, P. J., Vonrhein, C., et al. (2006) The crystal structure of iron-free human serum transferring provides insight into inter-lobe communication and receptor binding. J. Biol. Chem., in press.

    Google ScholarĀ 

  28. Kigawa, T., Yamaguchi-Nunokawa, E., Kodama, K., et al. (2002) Selenomethionine incorporation into a protein by cell-free synthesis. J. Struct. Funct. Genomics 2, 29ā€“35.

    ArticleĀ  CASĀ  PubMedĀ  Google ScholarĀ 

  29. Yokoyama, S. (2003) Protein expression systems for structural genomics and proteomics. Curr. Opin. Chem. Biol. 7, 39ā€“43.

    ArticleĀ  CASĀ  PubMedĀ  Google ScholarĀ 

  30. Hendrickson, W. A. and Ogata, C. M. (1997) Phase determination from multiwavelength anomalous diffraction measurements. Methods Enzymol. 276, 494ā€“523.

    ArticleĀ  CASĀ  Google ScholarĀ 

  31. Strub, M. P., Hoh, F., Sanchez, J. F., et al. (2003) Selenomethionine and selenocysteine double labeling strategy for crystallographic phasing. Structure (Camb) 11, 1359ā€“1367.

    ArticleĀ  CASĀ  Google ScholarĀ 

  32. Smith, J. L. and Thompson, A. (1998) Reactivity of selenomethionine: dents in the magic bullet? Structure 6, 815ā€“819.

    ArticleĀ  CASĀ  PubMedĀ  Google ScholarĀ 

  33. Sharff, A. J., Koronakis, E., Luisi, B., and Koronakis, V. (2000) Oxidation of selenomethionine: some MADness in the method! Acta Crystallogr. D Biol. Crystallogr. 56, 785ā€“788.

    ArticleĀ  CASĀ  PubMedĀ  Google ScholarĀ 

  34. Thomazeau, K., Curien, G., Thompson, A., Dumas, R., and Biou, V. (2001) MAD on threonine synthase: the phasing power of oxidized selenomethionine. Acta Crystallogr. D Biol. Crystallogr. 57, 1337ā€“1340.

    ArticleĀ  CASĀ  PubMedĀ  Google ScholarĀ 

  35. Boggon, T. J. and Shapiro, L. (2000) Screening for phasing atoms in protein crystallography. Structure Fold. Des. 8, R143ā€“R149.

    ArticleĀ  CASĀ  PubMedĀ  Google ScholarĀ 

  36. Leahy, D. J., Erickson, H. P., Aukhil, I., Joshi, P., and Hendrickson, W. A. (1994) Crystallization of a fragment of human fibronectin: introduction of methionine by site-directed mutagenesis to allow phasing via selenomethionine. Proteins 19, 48ā€“54.

    ArticleĀ  CASĀ  PubMedĀ  Google ScholarĀ 

  37. Skinner, M. M., Zhang, H., Leschnitzer, D. H., et al. (1994) Structure of the gene V protein of bacteriophage f1 determined by multiwavelength X-ray diffraction on the selenomethionyl protein. Proc. Natl. Acad. Sci. USA 91, 2071ā€“2075.

    ArticleĀ  CASĀ  PubMedĀ  Google ScholarĀ 

  38. Jones, D. T., Taylor, W. R., and Thornton, J. M. (1992) The rapid generation of mutation data matrices from protein sequences. Comput. Appl. Biosci. 8, 275ā€“282.

    CASĀ  PubMedĀ  Google ScholarĀ 

  39. Cohen, S. L. and Chait, B. T. (2001) Mass spectrometry as a tool for protein crystallography. Annu. Rev. Biophys. Biomol. Struct. 30, 67ā€“85.

    ArticleĀ  CASĀ  PubMedĀ  Google ScholarĀ 

  40. Hagemeier, C. H., Shima, S., Warkentin, E., Thauer, R. K., and Ermler, U. (2003) Coenzyme F420-dependent methylenetetrahydromethanopterin dehydrogenase from Methanopyrus kandleri: the selenomethionine-labelled and non-labelled enzyme crystallized in two different forms. Acta Crystallogr. D Biol. Crystallogr. 59, 1653ā€“1655.

    ArticleĀ  PubMedĀ  Google ScholarĀ 

  41. McAuley, K. E., Cummins, I., Papiz, M., Edwards, R., and Fordham-Skelton, A. P. (2003). Purification, crystallization and preliminary X-ray diffraction analysis of S-formylglutathione hydrolase from Arabidopsis thaliana: effects of pressure and selenomethionine substitution on space-group changes. Acta Crystallogr. D Biol. Crystallogr. 59, 2272ā€“2274.

    ArticleĀ  PubMedĀ  Google ScholarĀ 

  42. Poulsen, J. C., Harris, P., Jensen, K. F., and Larsen, S. (2001) Selenomethionine substitution of orotidine-5ā€²-monophosphate decarboxylase causes a change in crystal contacts and space group. Acta Crystallogr. D Biol. Crystallogr. 57, 1251ā€“1259.

    ArticleĀ  CASĀ  PubMedĀ  Google ScholarĀ 

  43. Witkowski, A. and Smith, S. (1998) Successful expression of a selenomethionyl protein under control of the temperature-sensitive lambda repressor requires higher than normal temperature. Biotechniques 24, 934ā€“936.

    CASĀ  PubMedĀ  Google ScholarĀ 

  44. Dong, C., Kotzsch, A., Dorward, M., van Pee, K. H., and Naismith, J. H. (2004) Crystallization and X-ray diffraction of a halogenating enzyme, tryptophan 7-halogenase, from Pseudomonas fluorescens. Acta Crystallogr. D Biol. Crystallogr. 60, 1438ā€“1440.

    ArticleĀ  PubMedĀ  Google ScholarĀ 

  45. Millard, C. S., Stols, L., Quartey, P., Kim, Y., Dementieva, I., and Donnelly, M. I. (2003) A less laborious approach to the high-throughput production of recombinant proteins in Escherichia coli using 2-liter plastic bottles. Protein Expr. Purif. 29, 311ā€“320.

    PubMedĀ  Google ScholarĀ 

  46. Stols, L., Millard, C. S., Dementieva, I., and Donnelly, M. I. (2004) Production of selenomethionine-labeled proteins in two-liter plastic bottles for structure determination. J. Struct. Funct. Genomics 5, 95ā€“102.

    ArticleĀ  CASĀ  PubMedĀ  Google ScholarĀ 

  47. Oā€™Reilly, D. R. and Miller, L. K. (1988) Expression and complex formation of simian virus 40 large T antigen and mouse p53 in insect cells. J. Virol. 62, 3109ā€“3119.

    PubMedĀ  Google ScholarĀ 

  48. Stein, A. J., Fuchs, G., Fu, C., Wolin, S. L., and Reinisch, K. M. (2005) Structural insights into RNA quality control: the Ro autoantigen binds misfolded RNAs via its central cavity. Cell 121, 529ā€“539.

    ArticleĀ  CASĀ  PubMedĀ  Google ScholarĀ 

  49. Gassner, N. C. and Matthews, B. W. (1999) Use of differentially substituted selenomethionine proteins in X-ray structure determination. Acta Crystallogr. D Biol. Crystallogr. 55, 1967ā€“1970.

    ArticleĀ  CASĀ  PubMedĀ  Google ScholarĀ 

  50. Bateman, K. S., Brownie, E. R., Wolodko, W. T., and Fraser, M. E. (2002) Structure of the mammalian CoA transferase from pig heart. Biochemistry 41, 14,455ā€“14,462.

    ArticleĀ  CASĀ  PubMedĀ  Google ScholarĀ 

  51. Carfi, A., Gong, H., Lou, H., et al. (2002) Crystallization and preliminary diffraction studies of the ectodomain of the envelope glycoprotein D from herpes simplex virus 1 alone and in complex with the ectodomain of the human receptor HveA. Acta Crystallogr. D Biol. Crystallogr. 58, 836ā€“838.

    ArticleĀ  PubMedĀ  Google ScholarĀ 

  52. Chen, W. Y. and Bahl, O. P. (1991) Selenomethionyl analog of recombinant human choriogonadotropin. J. Biol. Chem. 266, 9355ā€“9358.

    CASĀ  PubMedĀ  Google ScholarĀ 

  53. Liemann, S., Chandran, K., Baker, T. S., Nibert, M. L., and Harrison, S. C. (2002) Structure of the reovirus membrane-penetration protein, Mu1, in a complex with is protector protein, Sigma3. Cell 108, 283ā€“295.

    ArticleĀ  CASĀ  PubMedĀ  Google ScholarĀ 

  54. Fremont, D. H., Crawford, F., Marrack, P., Hendrickson, W. A., and Kappler, J. (1998) Crystal structure of mouse H2-M. Immunity 9, 385ā€“393.

    ArticleĀ  CASĀ  PubMedĀ  Google ScholarĀ 

Download references

Author information

Authors and Affiliations

Authors

Editor information

Editors and Affiliations

Rights and permissions

Reprints and permissions

Copyright information

Ā© 2007 Humana Press Inc.

About this protocol

Cite this protocol

DoubliƩ, S. (2007). Production of Selenomethionyl Proteins in Prokaryotic and Eukaryotic Expression Systems. In: Walker, J.M., DoubliƩ, S. (eds) Macromolecular Crystallography Protocols. Methods in Molecular Biology, vol 363. Humana Press. https://doi.org/10.1007/978-1-59745-209-0_5

Download citation

  • DOI: https://doi.org/10.1007/978-1-59745-209-0_5

  • Publisher Name: Humana Press

  • Print ISBN: 978-1-58829-292-6

  • Online ISBN: 978-1-59745-209-0

  • eBook Packages: Springer Protocols

Publish with us

Policies and ethics