A thermodynamic analysis of the sequence-specific binding of RNA by bacteriophage MS2 coat protein.
Most mutations in the sequence of the RNA hairpin that specifically binds MS2 coat protein either reduce the binding affinity or have no effect. However, one RNA mutation, a uracil to cytosine change in the loop, has the unusual property of increasing the binding affinity to the protein by nearly 100-fold. Guided by the structure of the protein-RNA complex, we used a series of protein mutations and RNA modifications to evaluate the thermodynamic basis for the improved affinity: The tight binding of the cytosine mutation is due to (i) the amino group of the cytosine residue making an intra-RNA hydrogen bond that increases the propensity of the free RNA to adopt the structure seen in the complex and (ii) the increased affinity of hydrogen bonds between the protein and a phosphate two bases away from the cytosine residue. The data are in good agreement with a recent comparison of the cocrystal structures of the two complexes, where small differences in the two structures are seen at the thermodynamically important sites.
Johansson
H E
HE
Department of Chemistry and Biochemistry, University of Colorado, Boulder, CO 80309-0215, USA.
Dertinger
D
D
LeCuyer
K A
KA
Behlen
L S
LS
Greef
C H
CH
Uhlenbeck
O C
OC
D001665:Binding Sites; D002213:Capsid / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D036022:Capsid Proteins; D016297:Mutagenesis, Site-Directed; D009690:Nucleic Acid Conformation; D011487:Protein Conformation; D012313:RNA / Q000737:chemistry / Q000378:metabolism*; D016601:RNA-Binding Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D013816:Thermodynamics
D036022:Capsid Proteins; D016601:RNA-Binding Proteins; D012313:RNA