PDBsum entry 1hza

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protein Protein-protein interface(s) links
Transcription PDB id
Protein chains
67 a.a. *
Waters ×110
* Residue conservation analysis
PDB id:
Name: Transcription
Title: Bacillus caldolyticus cold-shock protein mutants to study determinants of protein stability
Structure: Cold shock protein cspb. Chain: a, b. Synonym: cspb, bc-csp. Engineered: yes. Mutation: yes
Source: Bacillus caldolyticus. Organism_taxid: 1394. Gene: cspb. Expressed in: escherichia coli. Expression_system_taxid: 562
1.80Å     R-factor:   0.206     R-free:   0.246
Authors: H.Delbrueck,U.Mueller,D.Perl,F.X.Schmid,U.Heinemann
Key ref:
H.Delbrück et al. (2001). Crystal structures of mutant forms of the Bacillus caldolyticus cold shock protein differing in thermal stability. J Mol Biol, 313, 359-369. PubMed id: 11800562 DOI: 10.1006/jmbi.2001.5051
24-Jan-01     Release date:   07-Nov-01    
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Protein chains
Pfam   ArchSchema ?
P41016  (CSPB_BACCL) -  Cold shock protein CspB
66 a.a.
67 a.a.*
Key:    PfamA domain  Secondary structure  CATH domain
* PDB and UniProt seqs differ at 2 residue positions (black crosses)

 Gene Ontology (GO) functional annotation 
  GO annot!
  Cellular component     cytoplasm   1 term 
  Biological process     response to stress   3 terms 
  Biochemical function     nucleic acid binding     2 terms  


DOI no: 10.1006/jmbi.2001.5051 J Mol Biol 313:359-369 (2001)
PubMed id: 11800562  
Crystal structures of mutant forms of the Bacillus caldolyticus cold shock protein differing in thermal stability.
H.Delbrück, U.Mueller, D.Perl, F.X.Schmid, U.Heinemann.
The cold shock proteins Bc-Csp from the thermophile Bacillus caldolyticus and Bs-CspB from the mesophile Bacillus subtilis differ significantly in their conformational stability, although the two proteins differ by only 12 out of 67 amino acid residues. The three-dimensional structure of these small and compact beta-barrel proteins without disulfide bonds, cis-proline residues or tightly bound cofactors is very similar. Previous work has shown that Bc-Csp displays a twofold increase in the free energy of stabilization relative to its homolog Bs-CspB, and indicated that electrostatic interactions are, in part, responsible for this effect. It was further described that the stability difference is almost exclusively due to surface-exposed charged residues at sequence positions 3 and 66 of Bc-Csp and Bs-CspB, whereas all other amino acid changes between both proteins have no net effect on stability. To investigate how two surface residues determine the stability of Bc-Csp, Arg3 and Leu66 were replaced by glutamic acid, corresponding to the Bs-CspB sequence. The crystal structures of the resultant protein variants, Bc-Csp R3E and Bc-Csp L66E, were determined at 1.4 A and 1.27 A resolution, and refined to R values of 13.9 % and 15.8 %, respectively. Both structures closely resemble Bc-Csp in their global fold and show different hydrogen bonding and salt-bridge patterns when two independent molecules in the asymmetric unit of the crystal are compared. To extend the study to neighbored residues that help determine the surface charge around Arg3 and Leu66, the mutant proteins Bc-Csp E46A, Bc-Csp R3E/E46A/L66E and Bc-Csp V64T/L66E/67A were crystallized. Their structures were determined at resolutions of 1.8 A, 1.32 A and 1.8 A and refined to R values of 18.5 %, 13.8 % and 19.3 %, respectively. A systematic comparison of the crystal structures of all forms of the B. caldolyticus cold shock protein shows varying patterns of hydrogen bonds and electrostatic interactions around residues 3 and 66. Thermal destabilization of the protein by mutation appears to correlate with the extent of an acidic surface patch near the C-terminal carboxylate group.
  Selected figure(s)  
Figure 1.
Figure 1. Three-dimensional structure and sequence of the B. caldolyticus cold shock protein, Bc-Csp. The drawing shows the structure of Bc-Csp in two orientations (top). The residues mutated in this study cluster in one area of the protein surface and are labeled. In the right panel, the b-strands b1 to b5 and the connecting loops L1 to L4 are labeled as well. The amino acid sequence of Bc-Csp is compared with the sequence of the cold shock protein from B. subtilis, Bs-CspB (bottom). Sequence differences are highlighted in color. Mutations in residues marked red have little influence on protein stability[10]. Residues marked green are mutated in the crystal structures reported here. The residues mutated in Bc-Csp R3E/E46A/L66E or in Bc-Csp V64T/L66E/67A are printed in italics or underlined, respectively.
Figure 4.
Figure 4. Distribution of surface charges in Bc-Csp and its mutated variants. The molecules are oriented with the surface area bearing the mutations facing forward. A drawing of Bc-Csp is shown in the same orientation for comparison.
  The above figures are reprinted by permission from Elsevier: J Mol Biol (2001, 313, 359-369) copyright 2001.  
  Figures were selected by the author.  

Literature references that cite this PDB file's key reference

  PubMed id Reference
17964951 F.Dong, B.Olsen, and N.A.Baker (2008).
Computational methods for biomolecular electrostatics.
  Methods Cell Biol, 84, 843-870.  
17266726 K.E.Max, M.Zeeb, R.Bienert, J.Balbach, and U.Heinemann (2007).
Common mode of DNA binding to cold shock domains. Crystal structure of hexathymidine bound to the domain-swapped form of a major cold shock protein from Bacillus caldolyticus.
  FEBS J, 274, 1265-1279.
PDB code: 2hax
16698786 S.L.Moors, M.Hellings, M.De Maeyer, Y.Engelborghs, and A.Ceulemans (2006).
Tryptophan rotamers as evidenced by X-ray, fluorescence lifetimes, and molecular dynamics modeling.
  Biophys J, 91, 816-823.  
16844745 X.Huang, and H.X.Zhou (2006).
Similarity and difference in the unfolding of thermophilic and mesophilic cold shock proteins studied by molecular dynamics simulations.
  Biophys J, 91, 2451-2463.  
12923178 E.J.Stollar, U.Mayor, S.C.Lovell, L.Federici, S.M.Freund, A.R.Fersht, and B.F.Luisi (2003).
Crystal structures of engrailed homeodomain mutants: implications for stability and dynamics.
  J Biol Chem, 278, 43699-43708.
PDB codes: 1p7i 1p7j
14579351 G.Morra, M.Hodoscek, and E.W.Knapp (2003).
Unfolding of the cold shock protein studied with biased molecular dynamics.
  Proteins, 53, 597-606.  
12668430 H.X.Zhou, and F.Dong (2003).
Electrostatic contributions to the stability of a thermophilic cold shock protein.
  Biophys J, 84, 2216-2222.  
12943843 J.K.Yano, and T.L.Poulos (2003).
New understandings of thermostable and peizostable enzymes.
  Curr Opin Biotechnol, 14, 360-365.  
12843403 J.L.England, B.E.Shakhnovich, and E.I.Shakhnovich (2003).
Natural selection of more designable folds: a mechanism for thermophilic adaptation.
  Proc Natl Acad Sci U S A, 100, 8727-8731.  
12824494 K.B.Wong, C.F.Lee, S.H.Chan, T.Y.Leung, Y.W.Chen, and M.Bycroft (2003).
Solution structure and thermal stability of ribosomal protein L30e from hyperthermophilic archaeon Thermococcus celer.
  Protein Sci, 12, 1483-1495.
PDB codes: 1go0 1go1
12323355 B.van den Burg, and V.G.Eijsink (2002).
Selection of mutations for increased protein stability.
  Curr Opin Biotechnol, 13, 333-337.  
12496083 H.X.Zhou (2002).
Toward the physical basis of thermophilic proteins: linking of enriched polar interactions and reduced heat capacity of unfolding.
  Biophys J, 83, 3126-3133.  
The most recent references are shown first. Citation data come partly from CiteXplore and partly from an automated harvesting procedure. Note that this is likely to be only a partial list as not all journals are covered by either method. However, we are continually building up the citation data so more and more references will be included with time. Where a reference describes a PDB structure, the PDB code is shown on the right.