TY - JOUR
T1 - Specificity of Mnt 'master residue' obtained from in vivo and in vitro selections
AU - Silbaq, Fauzi S.
AU - Ruttenberg, Steven E.
AU - Stormo, Gary D.
N1 - Funding Information:
The authors thank Dr Stephen Elledge and Dr Ronald Davis for their donation of the transcriptional interference plasmids pNN388 and pNN396. We also thank Dr Kendall Knight and Dr Robert Sauer for their donation of Mnt plasmid pTM201, and Dr Larry Gold for the donation of E.coli strain JM107. We thank Nexagen for the gift of some DNA oligos. This work was supported by NIH grant GM28755.
PY - 2002/12/15
Y1 - 2002/12/15
N2 - Mnt is a repressor from phage P22 that belongs to the ribbon-helix-helix family of DNA binding factors. Four amino acids from the N-terminus of the protein, Arg2, His6, Asn8 and Arg10, interact with the base pairs of the DNA to provide the sequence specificity. Raumann et al. (Nature Struct. Biol., 2, 1115-1122) identified position 6 as a 'master residue' that controls the specificity of the protein. Models for the interaction have residue 6 of Mnt interacting directly with position 5 of the operator. In vivo selections demonstrated that protein variants at residue 6 bound specifically to operator mutations at that position. Operators in which the wild-type G at position 5 was replaced by T specifically bound to several different protein variants, primarily hydrophobic residues. The obtained protein variants, plus some others, were used in in vitro selections to determine their preferred binding sites. The results showed that the residue at position 6 influenced the preference for binding site bases predominantly at position 5, but that the effects of altering it can extend over longer distances, consistent with its designation as a 'master residue'. The similarities of binding sites for different residues do not correlate strongly with common measures of amino acid similarities.
AB - Mnt is a repressor from phage P22 that belongs to the ribbon-helix-helix family of DNA binding factors. Four amino acids from the N-terminus of the protein, Arg2, His6, Asn8 and Arg10, interact with the base pairs of the DNA to provide the sequence specificity. Raumann et al. (Nature Struct. Biol., 2, 1115-1122) identified position 6 as a 'master residue' that controls the specificity of the protein. Models for the interaction have residue 6 of Mnt interacting directly with position 5 of the operator. In vivo selections demonstrated that protein variants at residue 6 bound specifically to operator mutations at that position. Operators in which the wild-type G at position 5 was replaced by T specifically bound to several different protein variants, primarily hydrophobic residues. The obtained protein variants, plus some others, were used in in vitro selections to determine their preferred binding sites. The results showed that the residue at position 6 influenced the preference for binding site bases predominantly at position 5, but that the effects of altering it can extend over longer distances, consistent with its designation as a 'master residue'. The similarities of binding sites for different residues do not correlate strongly with common measures of amino acid similarities.
UR - http://www.scopus.com/inward/record.url?scp=0037115830&partnerID=8YFLogxK
U2 - 10.1093/nar/gkf684
DO - 10.1093/nar/gkf684
M3 - Review article
C2 - 12490722
AN - SCOPUS:0037115830
SN - 0305-1048
VL - 30
SP - 5539
EP - 5548
JO - Nucleic acids research
JF - Nucleic acids research
IS - 24
ER -