Thermal stability and atomic-resolution crystal structure of the Bacillus caldolyticus cold shock protein.
The bacterial cold shock proteins are small compact beta-barrel proteins without disulfide bonds, cis-proline residues or tightly bound cofactors. Bc-Csp, the cold shock protein from the thermophile Bacillus caldolyticus shows a twofold increase in the free energy of stabilization relative to its homolog Bs-CspB from the mesophile Bacillus subtilis, although the two proteins differ by only 12 out of 67 amino acid residues. This pair of cold shock proteins thus represents a good system to study the atomic determinants of protein thermostability. Bs-CspB and Bc-Csp both unfold reversibly in cooperative transitions with T(M) values of 49.0 degrees C and 77.3 degrees C, respectively, at pH 7.0. Addition of 0.5 M salt stabilizes Bs-CspB but destabilizes Bc-Csp. To understand these differences at the structural level, the crystal structure of Bc-Csp was determined at 1.17 A resolution and refined to R=12.5% (R(free)=17.9%). The molecular structures of Bc-Csp and Bs-CspB are virtually identical in the central beta-sheet and in the binding region for nucleic acids. Significant differences are found in the distribution of surface charges including a sodium ion binding site present in Bc-Csp, which was not observed in the crystal structure of the Bs-CspB. Electrostatic interactions are overall favorable for Bc-Csp, but unfavorable for Bs-CspB. They provide the major source for the increased thermostability of Bc-Csp. This can be explained based on the atomic-resolution crystal structure of Bc-Csp. It identifies a number of potentially stabilizing ionic interactions including a cation-binding site and reveals significant changes in the electrostatic surface potential.
Mueller
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Forschungsgruppe Kristallographie, Max-Delbrück-Centrum für Molekulare Medizin, Robert-Roessle-Str. 10, Berlin, D-13125, Germany.
Perl
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Schmid
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Heinemann
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D000595:Amino Acid Sequence; D001407:Bacillus / Q000737:chemistry*; D001426:Bacterial Proteins / Q000737:chemistry*; D001665:Binding Sites; D002412:Cations / Q000378:metabolism / Q000494:pharmacology; D002942:Circular Dichroism; D018360:Crystallography, X-Ray; D006360:Heat-Shock Proteins / Q000737:chemistry* / Q000378:metabolism*; D006358:Hot Temperature; D008958:Models, Molecular; D008969:Molecular Sequence Data; D009696:Nucleic Acids / Q000378:metabolism; D009994:Osmolar Concentration; D011489:Protein Denaturation / Q000187:drug effects; D017510:Protein Folding; D017433:Protein Structure, Secondary / Q000187:drug effects; D017434:Protein Structure, Tertiary / Q000187:drug effects; D012964:Sodium / Q000378:metabolism / Q000494:pharmacology; D055672:Static Electricity; D013329:Structure-Activity Relationship; D013816:Thermodynamics
D001426:Bacterial Proteins; D002412:Cations; D006360:Heat-Shock Proteins; D009696:Nucleic Acids; C407094:cold shock protein, Bacillus; C077188:cold-shock protein CspB, Bacteria; D012964:Sodium