Solution conformations of DNAs containing binding sites of the catabolite gene activator protein and lac repressor protein: characterization by Raman spectroscopy.
Raman spectra from three subfragments of the Escherichia coli lactose promoter region were obtained in 0.1 M NaCl. The three DNAs are 21, 40, and 62 bp in length. The 21 and 62 bp DNAs contain the binding site for the catabolite gene activator protein (CAP). The 40 bp DNA contains the binding site for the lac repressor. A quantitative analysis of Raman band characteristics indicates an overall B-type conformation for these gene regulatory sites. Bands which correspond to A-family (807 cm-1) and B-family (834 cm-1) deoxyribose phosphate vibrations have the same intensities as bands found in heterogeneous DNAs. The spectra of the 21 bp CAP site have, however, a small band at 867 cm-1 and several other small differences similar to some characteristics observed in C-DNA spectra. Several dG nucleosides in the CAP site appear to be altered from the conventional C2'-endo/anti conformation. At 45 degrees C, well below the melting region of these DNAs, small changes occur in the spectra of the 40 bp lac repressor site which are not observed in the other DNAs. A weak band occurs at 705 cm-1, and intensity changes are observed at 497, 682, and 792 cm-1. The changes suggest that the conformations of several dG nucleosides are altered and that a small region may exist with characteristics of an A-family backbone. This conformational change at 45 degrees C coincides with previous NMR observations indicating an enhanced imino proton exchange rate at a GTG sequence within the lac operator site.
DeGrazia
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School of Physics, Georgia Institute of Technology, Atlanta 30332.
Brown
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D
Cheung
S
S
Wartell
R M
RM
D001665:Binding Sites; D002373:Cyclic AMP Receptor Protein / Q000378:metabolism*; D004269:DNA, Bacterial / Q000235:genetics / Q000378:metabolism* / Q000648:ultrastructure; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism*; D005798:Genes, Bacterial; D005809:Genes, Regulator; D009690:Nucleic Acid Conformation; D011401:Promoter Regions, Genetic; D011485:Protein Binding; D012097:Repressor Proteins / Q000378:metabolism*; D012996:Solutions; D013059:Spectrum Analysis, Raman / Q000379:methods; D014157:Transcription Factors / Q000378:metabolism*
D002373:Cyclic AMP Receptor Protein; D004269:DNA, Bacterial; D012097:Repressor Proteins; D012996:Solutions; D014157:Transcription Factors