Reprogramming EF-hands for design of catalytically amplified lanthanide sensors

  • Korrie L. Mack
  • Olesia V. Moroz
  • Yurii S. Moroz
  • Alissa B. Olsen
  • Jaclyn M. McLaughlin
  • Ivan V. Korendovych
Original Paper

Abstract

We recently reported that a computationally designed catalyst nicknamed AlleyCat facilitates C–H proton abstraction in Kemp elimination at neutral pH in a selective and calcium-dependent fashion by a factor of approximately 100,000 (Korendovych et al. in Proc. Natl. Acad. Sci. USA 108:6823, 2011). Kemp elimination produced a colored product that can be easily read out, thus making AlleyCat a catalytically amplified metal sensor for calcium. Here we report that metal-binding EF-hand motifs in AlleyCat could be redesigned to incorporate trivalent metal ions without significant loss of catalytic activity. Mutation of a single neutral residue at position 9 of each of the EF-hands to glutamate results in almost a two orders of magnitude improvement of selectivity for trivalent metal ions over calcium. Development of this new lanthanide-dependent switchable Kemp eliminase, named CuSeCat EE, provides the foundation for further selectivity improvement and broadening the scope of the repertoire of metals for sensing. A concerted effort in the design of switchable enzymes has many environmental, sensing, and metal ion tracking applications.

Keywords

Protein design Metal sensor Lanthanides 

Supplementary material

775_2013_985_MOESM1_ESM.pdf (5.1 mb)
Supplementary material 1 (PDF 5217 kb)

References

  1. 1.
    Domaille DW, Que EL, Chang CJ (2008) Nat Chem Biol 4:168–175PubMedCrossRefGoogle Scholar
  2. 2.
    Luque de Castro MD, Herrera MC (2003) Biosens Bioelectron 18:279–294Google Scholar
  3. 3.
    Doi N, Yanagawa H (1999) FEBS Lett 453:305–307Google Scholar
  4. 4.
    Dwyer MA, Hellinga HW (2004) Curr Opin Chem Biol 14:495–504Google Scholar
  5. 5.
    Miyawaki A, Llopis J, Heim R, McCaffery JM, Adams JA, Ikura M, Tsien RY (1997) Nature 388:882–887Google Scholar
  6. 6.
    Zhu L, Anslyn E (2006) Angew Chem Int Ed 45:1190–1196CrossRefGoogle Scholar
  7. 7.
    Ostermeier M (2009) Curr Opin Struct Biol 19:442–448PubMedCrossRefGoogle Scholar
  8. 8.
    Yoon HJ, Kuwabara J, Kim J-H, Mirkin CA (2010) Science 330:66–69PubMedCrossRefGoogle Scholar
  9. 9.
    Korendovych IV, Kulp DW, Wu Y, Cheng H, Roder H, DeGrado WF (2011) Proc Natl Acad Sci USA 108:6823–6827PubMedCrossRefGoogle Scholar
  10. 10.
    Blommel PG, Fox BG (2007) Protein Expr Purif 55:53–68PubMedCrossRefGoogle Scholar
  11. 11.
    Studier FW (2005) Protein Expr Purif 41:207–234PubMedCrossRefGoogle Scholar
  12. 12.
    Rothlisberger D, Khersonsky O, Wollacott AM, Jiang L, DeChancie J, Betker J, Gallaher JL, Althoff EA, Zanghellini A, Dym O, Albeck S, Houk KN, Tawfik DS, Baker D (2008) Nature 453:190–195PubMedCrossRefGoogle Scholar
  13. 13.
    Urbauer JL, Short JH, Dow LK, Wand AJ (1995) Biochemistry 34:8099–8109PubMedCrossRefGoogle Scholar
  14. 14.
    Radivoyevitch T (2009) Biol Direct 4:49PubMedCrossRefGoogle Scholar
  15. 15.
    Masino L, Martin SR, Bayley PM (2000) Protein Sci 9:1519–1529PubMedCrossRefGoogle Scholar
  16. 16.
    Marley J, Lu M, Bracken C (2001) J Biomol NMR 20:71–75PubMedCrossRefGoogle Scholar
  17. 17.
    Klee CB, Crouch TH, Richman PG (1980) Ann Rev Biochem 49:489–515PubMedCrossRefGoogle Scholar
  18. 18.
    Bertini I, Gelis I, Katsaros N, Luchinat C, Provenzani A (2003) Biochemistry 42:8011–8021PubMedCrossRefGoogle Scholar
  19. 19.
    Le Clainche L, Plancque G, Amekraz B, Moulin C, Pradines-Lecomte C, Peltier G, Vita C (2003) J Biol Inorg Chem 8:334–340PubMedGoogle Scholar
  20. 20.
    am Ende CW, Meng HY, Ye M, Pandey AK, Zondlo NJ (2010) ChemBioChem 11:1738–1747Google Scholar
  21. 21.
    Zondlo SC, Gao F, Zondlo NJ (2010) J Am Chem Soc 132:5619–5621PubMedCrossRefGoogle Scholar
  22. 22.
    Moroz OV, Moroz YS, Mack KL, Wu Y, Olsen AB, Cheng H, McLaughlin JM, Raymond EA, Roder H, Korendovych IV (in preparation)Google Scholar
  23. 23.
    Drake SK, Falke JJ (1996) Biochemistry 35:1753–1760PubMedCrossRefGoogle Scholar
  24. 24.
    Drake SK, Lee KL, Falke JJ (1996) Biochemistry 35:6697–6705PubMedCrossRefGoogle Scholar
  25. 25.
    Falke JJ, Drake SK, Hazard AL, Peersen OB (1994) Q Rev Biophys 27:219–290PubMedCrossRefGoogle Scholar
  26. 26.
    Ye YM, Lee H-W, Yang W, Shealy S, Yang JJ (2005) J Am Chem Soc 127:3743–3750PubMedCrossRefGoogle Scholar
  27. 27.
    Chou JJ, Li S, Klee CB, Bax A (2001) Nat Struct Biol 8:990–997PubMedCrossRefGoogle Scholar
  28. 28.
    O’Neil KT, DeGrado WF (1990) Trends Biochem Sci 15:59–64PubMedCrossRefGoogle Scholar
  29. 29.
    Snyder EE, Buoscio BW, Falke JJ (1990) Biochemistry 29:3937–3943PubMedCrossRefGoogle Scholar
  30. 30.
    Mulqueen P, Tingey JM, Horrocks WD Jr (1985) Biochemistry 24:6639–6645PubMedCrossRefGoogle Scholar
  31. 31.
    Samish I, MacDermaid CM, Perez-Aguilar JS, Saven JG (2011) Annu Rev Phys Chem 62:129–149PubMedCrossRefGoogle Scholar
  32. 32.
    DeGrado WF, Summa CM, Pavone V, Nastri F, Lombardi A (1999) Annu Rev Biochem 68:779–819PubMedCrossRefGoogle Scholar
  33. 33.
    Das R, Baker D (2008) Annu Rev Biochem 77:363–382PubMedCrossRefGoogle Scholar
  34. 34.
    Brustad EM, Arnold FH (2011) Curr Opin Chem Biol 15:201–210PubMedCrossRefGoogle Scholar
  35. 35.
    Nitz M, Franz KJ, Maglathlin RL, Imperiali B (2003) ChemBioChem 4:272–276PubMedCrossRefGoogle Scholar
  36. 36.
    Franz KJ, Nitz M, Imperiali B (2003) ChemBioChem 4:265–271PubMedCrossRefGoogle Scholar
  37. 37.
    Nitz M, Sherawat M, Franz KJ, Peisach E, Allen KN, Imperiali B (2004) Angew Chem Int Ed 43:3682–3685Google Scholar

Copyright information

© SBIC 2013

Authors and Affiliations

  • Korrie L. Mack
    • 1
  • Olesia V. Moroz
    • 1
  • Yurii S. Moroz
    • 1
  • Alissa B. Olsen
    • 1
  • Jaclyn M. McLaughlin
    • 1
  • Ivan V. Korendovych
    • 1
  1. 1.Department of ChemistrySyracuse UniversitySyracuseUSA

Personalised recommendations