Quantitative modeling of DNA-protein interactions: effects of amino acid substitutions on binding specificity of the Mnt repressor.
Understanding DNA-protein recognition quantitatively is essential to developing computational algorithms for accurate transcriptional binding site prediction. Using a quantitative, multiple fluorescence, relative affinity (QuMFRA) assay, we determine the binding specificity of 11 different position 6 variants of the Mnt repressor for operators containing all 16 possible dinucleotides at operator positions 16 and 17. We show that the wild-type and all variant proteins interact with the two positions in a non-independent manner, but that a simple independent model provides a close approximation to the true binding affinities. The wild-type His at amino acid 6 is the only protein to prefer the AC sequence of the wild-type operator, whereas most of the variant proteins prefer TA. H6R is unique in having a strong preference for C at position 16. A comparison of the quantitative binding data for all of the protein variants with a model for recognition of the early growth response (EGR) zinc finger family suggests that interactions of Mnt with positions 16 and 17 are similar to interactions of EGR with positions 1 and 2, respectively. This information leads to an augmented model for the interaction of Mnt with its operator.
Department of Genetics, Washington University, 660 S. Euclid, Box 8232, St Louis, MO 63110, USA.
Tsz-Kwong
TK
Man
Joshua SungWoo
JS
Yang
Gary D
GD
Stormo
D019943:Amino Acid Substitution; D001665:Binding Sites; D004247:DNA / Q000737:chemistry / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D008957:Models, Genetic; D009875:Operator Regions, Genetic; D012097:Repressor Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D014764:Viral Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D054334:Viral Regulatory and Accessory Proteins
D004268:DNA-Binding Proteins; D012097:Repressor Proteins; D014764:Viral Proteins; D054334:Viral Regulatory and Accessory Proteins; C016649:phage repressor proteins; D004247:DNA