Skip to main content

Thank you for visiting nature.com. You are using a browser version with limited support for CSS. To obtain the best experience, we recommend you use a more up to date browser (or turn off compatibility mode in Internet Explorer). In the meantime, to ensure continued support, we are displaying the site without styles and JavaScript.

Two exposed amino acid residues confer thermostability on a cold shock protein

Abstract

Thermophilic organisms produce proteins of exceptional stability. To understand protein thermostability at the molecular level we studied a pair of cold shock proteins, one of mesophilic and one of thermophilic origin, by systematic mutagenesis. Although the two proteins differ in sequence at 12 positions, two surface-exposed residues are responsible for the increase in stability of the thermophilic protein (by 15.8 kJ mol−1 at 70 °C). 11.5 kJ mol−1 originate from a predominantly electrostatic contribution of Arg 3 and 5.2 kJ mol−1 from hydrophobic interactions of Leu 66 at the carboxy terminus. The mesophilic protein could be converted to a highly thermostable form by changing the Glu residues at positions 3 and 66 to Arg and Leu, respectively. The variation of surface residues may thus provide a simple and powerful approach for increasing the thermostability of a protein.

This is a preview of subscription content

Access options

Rent or Buy article

Get time limited or full article access on ReadCube.

from$8.99

All prices are NET prices.

Figure 1: The mesophilic cold shock protein Bs-CspB differs from its thermophilic homolog, Bc-Csp, at 12 sequence positions.
Figure 2: Thermal stability curves.
Figure 3: Effect of all individual sequence differences on the stability of Bc-Csp.
Figure 4: The residues that contribute most to the difference in thermal stability between Bs-CspB and Bc-Csp are located near each other at the protein surface.

References

  1. 1

    Jaenicke, R. & Böhm, G. Curr. Opin. Struct. Biol. 8, 738–748 (1998).

    CAS  Article  Google Scholar 

  2. 2

    Makhatadze, G.I. & Privalov, P.L. Adv. Protein Chem. 47, 307–425 ( 1995).

    CAS  Article  Google Scholar 

  3. 3

    Graumann, P.L. & Marahiel, M.A. Trends Biochem. Sci. 23, 286–290 (1998).

    CAS  Article  Google Scholar 

  4. 4

    Brandi, A., Spurio, R., Gualerzi, C.O. & Pon, C.L. EMBO J. 18, 1653–1659 ( 1999).

    CAS  Article  Google Scholar 

  5. 5

    Schindelin, H., Marahiel, M.A. & Heinemann, U. Nature 364, 164– 168 (1993).

    CAS  Article  Google Scholar 

  6. 6

    Müller, U., Perl, D., Schmid, F.X. & Heinemann, U. J. Mol. Biol. 297, 975–988 ( 2000).

    Article  Google Scholar 

  7. 7

    Perl, D., Welker, C., Schindler, T., Schröder, K., Marahiel, M.A., Jaenicke, R. & Schmid, F.X. Nature Struct. Biol. 5, 229–235 (1998).

    CAS  Article  Google Scholar 

  8. 8

    Mayr, L.M., Landt, O., Hahn, U. & Schmid, F.X. J. Mol. Biol. 231, 897–912 ( 1993).

    CAS  Article  Google Scholar 

  9. 9

    Grimsley, G.R. et al. Protein Sci. 8, 1843– 1849 (1999).

    CAS  Article  Google Scholar 

  10. 10

    Loladze, V.V., Ibarra-Molero, B., Sanchez-Ruiz, J.M. & Makhatadze, G.I. Biochemistry 38, 16419–16423 (1999).

    CAS  Article  Google Scholar 

  11. 11

    Spector, S. et al. Biochemistry 39, 872– 879 (2000).

    CAS  Article  Google Scholar 

  12. 12

    Karshikoff, A. & Ladenstein, R. Protein Eng. 11, 867–872 ( 1998).

    CAS  Article  Google Scholar 

  13. 13

    Lebbink, J.H., Knapp, S., van der Oost, J., Rice, D., Ladenstein, R. & de Vos, W.M. J. Mol. Biol. 280, 287– 296 (1998).

    CAS  Article  Google Scholar 

  14. 14

    Lebbink, J.H., Knapp, S., van der Oost, J., Rice, D., Ladenstein, R. & de Vos, W.M. J. Mol. Biol. 289, 357– 369 (1999).

    CAS  Article  Google Scholar 

  15. 15

    Knapp, S., Kardinahl, S., Hellgren, N., Tibbelin, G., Schafer, G. & Ladenstein, R. J. Mol. Biol. 285, 689– 702 (1999).

    CAS  Article  Google Scholar 

  16. 16

    Macedo-Ribeiro, S., Darimont, B., Sterner, R. & Huber, R. Structure 4, 1291–1301 (1996).

    CAS  Article  Google Scholar 

  17. 17

    Kawamura, S., Abe, Y., Ueda, T., Masumoto, K., Imoto, T., Yamasaki, N. & Kimura, M. J. Biol. Chem. 273, 19982– 19987 (1998).

    CAS  Article  Google Scholar 

  18. 18

    Haney, P.J., Badger, J.H., Buldak, G.L., Reich, C.I., Woese, C.R. & Olsen, G.J. Proc. Natl. Acad. Sci. USA 96, 3578–3583 (1999).

    CAS  Article  Google Scholar 

  19. 19

    Elcock, A.H. J. Mol. Biol. 284, 489–502 (1998).

    CAS  Article  Google Scholar 

  20. 20

    Vetriani, C. et al. Proc. Natl. Acad. Sci. USA 95, 12300 –12305 (1998).

    CAS  Article  Google Scholar 

  21. 21

    de Bakker, P.I., Hunenberger, P.H. & McCammon, J.A. J. Mol. Biol. 285, 1811– 1830 (1999).

    CAS  Article  Google Scholar 

  22. 22

    Hendsch, Z.S. & Tidor, B. Protein Sci. 8, 1381–1392 (1999).

    CAS  Article  Google Scholar 

  23. 23

    Xiao, L. & Honig, B. J. Mol. Biol. 289, 1435–1444 (1999).

    CAS  Article  Google Scholar 

  24. 24

    Street, A.G. & Mayo, S.L. Structure 7, R105–R109 (1999).

    CAS  Article  Google Scholar 

  25. 25

    Desjarlais, J.R. & Handel, T.M. J. Mol. Biol. 290, 305–318 ( 1999).

    CAS  Article  Google Scholar 

  26. 26

    Sieber, V., Plückthun, A. & Schmid, F.X. Nature Biotechnol. 16, 955– 960 (1998).

    CAS  Article  Google Scholar 

  27. 27

    Arnold, F.A. & Volkov, A.A. Curr. Opin. Chem. Biol. 3, 54–59 (1999).

    CAS  Article  Google Scholar 

  28. 28

    Finucane, M.D., Tuna, M., Lees, J.H. & Woolfson, D.N. Biochemistry 38, 11604–11612 ( 1999).

    CAS  Article  Google Scholar 

  29. 29

    Willimsky, G., Bang, H., Fischer, G. & Marahiel, M.A. J. Bacteriol. 174, 6326–6335 ( 1992).

    CAS  Article  Google Scholar 

  30. 30

    Schindelin, H., Herrler, M., Willimsky, G., Marahiel, M.A. & Heinemann, U. Proteins 14, 120–124 (1992).

    CAS  Article  Google Scholar 

Download references

Acknowledgements

We thank the members of our laboratories and the Marahiel laboratory for help and discussions as well as C. Brooks III (Scripps Research Institute) and C. Nick Pace (Texas A&M University) for a fruitful exchange of ideas about the electrostatic stabilization of proteins. This work was supported by grants from the Deutsche Forschungsgemeinschaft and the Fonds der Chemischen Industrie.

Author information

Affiliations

Authors

Corresponding author

Correspondence to Franz X. Schmid.

Rights and permissions

Reprints and Permissions

About this article

Cite this article

Perl, D., Mueller, U., Heinemann, U. et al. Two exposed amino acid residues confer thermostability on a cold shock protein. Nat Struct Mol Biol 7, 380–383 (2000). https://doi.org/10.1038/75151

Download citation

Further reading

Search

Quick links

Nature Briefing

Sign up for the Nature Briefing newsletter — what matters in science, free to your inbox daily.

Get the most important science stories of the day, free in your inbox. Sign up for Nature Briefing