PDBsum entry 2es2

Gene regulation

PDB id: 2es2

Contents

Protein chain

67 a.a. *

DNA/RNA

Metals

_CA ×2

Waters ×63

* Residue conservation analysis

PDB id:

2es2

Name:

Gene regulation

Title:

Crystal structure analysis of the bacillus subtilis cold shock protein bs-cspb in complex with hexathymidine

Structure:

5'-d( Tp Tp Tp Tp Tp T)-3'. Chain: b. Synonym: hexathymidine. Engineered: yes. Cold shock protein cspb. Chain: a. Synonym: major cold shock protein. Engineered: yes

Source:

Synthetic: yes. Other_details: DNA synthesis. Bacillus subtilis. Organism_taxid: 1423. Gene: cspb, cspa. Expressed in: escherichia coli bl21(de3). Expression_system_taxid: 469008.

Biol. unit:

Dimer (from PQS)

Resolution:

1.78Å R-factor: 0.190 R-free: 0.223

Authors:

K.E.A.Max,M.Bienert,U.Heinemann

Key ref:

K.E.Max et al. (2006). T-rich DNA single strands bind to a preformed site on the bacterial cold shock protein Bs-CspB. J Mol Biol, 360, 702-714. PubMed id: 16780871 DOI: 10.1016/j.jmb.2006.05.044

Date:

25-Oct-05 Release date: 05-Sep-06

Protein chain

P32081  (CSPB_BACSU) -  Cold shock protein CspB from Bacillus subtilis (strain 168)

Seq: 67 a.a.

Struc: 67 a.a.

DNA/RNA chain

T-T-T-T-T-T 6 bases

DOI no: 10.1016/j.jmb.2006.05.044

J Mol Biol 360:702-714 (2006)

PubMed id: 16780871

T-rich DNA single strands bind to a preformed site on the bacterial cold shock protein Bs-CspB.

K.E.Max, M.Zeeb, R.Bienert, J.Balbach, U.Heinemann.

ABSTRACT

Bacterial cold shock proteins (CSPs) are involved in cellular adaptation to cold stress. They bind to single-stranded nucleic acids with a KD value in the micro- to nanomolar range. Here we present the structure of the Bacillus subtilis CspB (Bs-CspB) in complex with hexathymidine (dT6) at a resolution of 1.78 A. Bs-CspB binds to dT6 with nanomolar affinity via an amphipathic interface on the protein surface. Individual binding subsites interact with single nucleobases through stacking interactions and hydrogen bonding. The sugar-phosphate backbone and the methyl groups of the thymine nucleobases remain solvent exposed and are not contacted by protein groups. Fluorescence titration experiments monitoring the binding of oligopyrimidines to Bs-CspB reveal binding preferences at individual subsites and allow the design of an optimised heptapyrimidine ligand, which is bound with sub-nanomolar affinity. This study reveals the stoichiometry and sequence determinants of the binding of single-stranded nucleic acids to a preformed site on Bs-CspB and thus provides the structural basis of the RNA chaperone and transcription antitermination activities of the CSP.

Selected figure(s)

Figure 1.
Figure 1. Hexathymidine bound to Bs-CspB. (a) Topological representation of the Bs-CspB ligand binding site. Residues from two ligand molecules (dT[6]), which are associated with one Bs-CspB, are shown in beige and red (backbone). The DNA nucleotides and associated symmetry related protein molecules are rendered as semitransparent grey. (b) Electrostatic surface potential of the Bs-CspB ligand binding site. The potential is projected onto the contact surface of the protein. The ligand and surrounding are depicted as described for (a). (c) Continuous arrangement formed by protein (green) and dT[6] molecules (beige, backbone red). β-Strands from the protein are displayed as green arrows. All Figures were drawn with Pymol (http://pymol.sourceforge.net/), the electrostatic surface potential was calculated with APBS^35 for pH 7, with a range from −10 (red) to +10 kT (blue).

Figure 3.
Figure 3. Hydrophobic and polar interactions between dT[6] and Bs-CspB. (a) The contact surface of Bs-CspB is shown as a semitransparent grey object, protein groups involved in stacking interactions and hydrogen bonding are colored according to the CPK scheme with the exception of carbon which is in green. Hydrogen bonds between protein and DNA groups are depicted as dotted lines. This is a stereo picture. (b) Representative section from the continuous arrangement of DNA (black) interacting with Bs-CspB groups (grey), displayed as structural formulas (bb, protein backbone). All parts of a complete binding site from a single protein molecule are highlighted by a grey box. Stacking interactions between aromatic side-chains and nucleobases, originating from the centers (stack) or rims (edge-on stack) of the aromatic rings are depicted as solid grey lines. Interactions involving hydrogen bonds are displayed as dotted lines, water molecules mediating protein–ligand interactions are depicted as small spheres. Adjacent side-chains belong to symmetry related molecules. The numbers of the contact subsites for individual nucleobases are given at the bottom. These subsites are discussed in the text.

The above figures are reprinted by permission from Elsevier: J Mol Biol (2006, 360, 702-714) copyright 2006.

Figures were selected by the author.