1998
DOI: 10.1073/pnas.95.16.9244
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A thermodynamic analysis of the sequence-specific binding of RNA by bacteriophage MS2 coat protein

Abstract: 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 cytosin… Show more

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Cited by 92 publications
(88 citation statements)
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“…Other codon-directed mutagenesis experiments confirmed earlier results showing that the identity of Asn-87 was for specific binding of MS2 RNA and for discrimination of Qb RNA binding (Spingola and Peabody, 1997). Intensively were studied also crystal structures of complexes between recombinant MS2 capsids with mutations and RNA operator sequences (Valegârd et al, 1997;Johansson et al, 1998;Helgstrand et al, 2002;Horn et al, 2004). The conversion of Asn-87 to Ala improved the ability of purine-RNAs (GA operator type) to bind MS2 coat protein, but binding to pyrimidine-RNAs (MS2 operator type) still remained tighter (Johansson et al, 1998).…”
Section: Introductionsupporting
confidence: 68%
See 1 more Smart Citation
“…Other codon-directed mutagenesis experiments confirmed earlier results showing that the identity of Asn-87 was for specific binding of MS2 RNA and for discrimination of Qb RNA binding (Spingola and Peabody, 1997). Intensively were studied also crystal structures of complexes between recombinant MS2 capsids with mutations and RNA operator sequences (Valegârd et al, 1997;Johansson et al, 1998;Helgstrand et al, 2002;Horn et al, 2004). The conversion of Asn-87 to Ala improved the ability of purine-RNAs (GA operator type) to bind MS2 coat protein, but binding to pyrimidine-RNAs (MS2 operator type) still remained tighter (Johansson et al, 1998).…”
Section: Introductionsupporting
confidence: 68%
“…Intensively were studied also crystal structures of complexes between recombinant MS2 capsids with mutations and RNA operator sequences (Valegârd et al, 1997;Johansson et al, 1998;Helgstrand et al, 2002;Horn et al, 2004). The conversion of Asn-87 to Ala improved the ability of purine-RNAs (GA operator type) to bind MS2 coat protein, but binding to pyrimidine-RNAs (MS2 operator type) still remained tighter (Johansson et al, 1998). The authors also appointed the possible role of some other amino acids in the complex formation and particularly of Ser-52, Asn-55 and Lys-57.…”
Section: Introductionmentioning
confidence: 99%
“…We suspect that the MS2d-GFP proteins bound to the RNA molecule might ''immortalize'' it, i.e., prevent or at least considerably slow down its degradation. The protein͞RNA off-rate for the binding site sequence in use is on the order of many hours (52)(53)(54), which suggests that MS2d-GFP might not be displaced by the various RNA-degrading enzymes in the cell.…”
Section: Results and Discussion Ms2-rna Complexes In Vitromentioning
confidence: 99%
“…Pum binds to the PRE with a dissociation constant in the low nano-molar range (Zamore et al 1997(Zamore et al , 1999. To strengthen this interaction, we utilized a modified MS2 coat protein and binding site interaction with an order of magnitude stronger binding (Lim et al 1994;Johansson et al 1998). Two reporters were used in this analysis: RnLuc MS2 pA-the 3 ′ poly(A) tail was generated using efficient cleavage/polyadenylation elements, whereas in the RnLuc MS2 HSL reporter, a nonadenylated 3 ′ end was generated by the HSL (Fig.…”
Section: The Rbd Of Pumilio Enhances Deadenylation Dependent On the Pmentioning
confidence: 99%