T-rich DNA single strands bind to a preformed site on the bacterial cold shock protein Bs-CspB.
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.
Max
Klaas E A
KE
Max-Delbrück-Centrum für Molekulare Medizin 13125 Berlin, Germany.
Zeeb
Markus
M
Bienert
Ralf
R
Balbach
Jochen
J
Heinemann
Udo
U
D000595:Amino Acid Sequence; D001412:Bacillus subtilis / Q000235:genetics / Q000378:metabolism*; D001426:Bacterial Proteins / Q000737:chemistry* / Q000378:metabolism*; D001665:Binding Sites; D004277:DNA, Single-Stranded / Q000378:metabolism*; D006360:Heat-Shock Proteins / Q000737:chemistry* / Q000378:metabolism*; D057927:Hydrophobic and Hydrophilic Interactions; D008024:Ligands; D008958:Models, Molecular; D008969:Molecular Sequence Data; D009154:Mutation / Q000235:genetics; D009841:Oligonucleotides / Q000737:chemistry; D011485:Protein Binding; D017433:Protein Structure, Secondary; D013941:Thymine / Q000737:chemistry*
D001426:Bacterial Proteins; D004277:DNA, Single-Stranded; D006360:Heat-Shock Proteins; D008024:Ligands; D009841:Oligonucleotides; C407094:cold shock protein, Bacillus; D013941:Thymine