Analysis of DNA-Protein Interactions in Complexes of Transcription Factor NF-κB with DNA

Abstract

We have applied bioinformatic analysis of X-ray 3D structures of complexes of transcription factor NF-kappaB with DNAs. We determined the number of possible Van der Waals contacts and hydrogen bonds between amino acid residues and nucleotides. Conservative contacts in the NF-kappaB dimer-DNA complex composed of p50 and/or p65 NF-kappaB subunit and DNA sequences like 5 -GGGAMWTTCC-3 were revealed. Based on these results, we propose a novel scheme for interactions between NF-kappaB p50 homodimer and the kappaB region of the immunoglobulin light chain gene enhancer (Ig-kappaB). We applied a chemical cross-linking technique to study the proximity of some Lys and Cys residues of NF-kappaB p50 subunit with certain reactive nucleotides into its recognition site. In all cases, the experimentally determined protein-DNA contacts were in good agreement with the predicted ones.

Thymidine Glycol: The Effect on DNA Structure and DNA Binding by Site-Specific Proteins

“…As mentioned above (see also [48]) oxidation of thymidine residue (especially flanked by А/Т pairs from the both sides) results in Tg extrahelical localization and in the changed conformation of the 5'-phosphate group of the Tg nucleotide. A generalized scheme of interaction between р50/р50 dimer and its target DNA [66] suggests that maximal number of the DNA protein contacts should be broken when Tg is incorporated in the position 16 of the κB site, as found here.…”

Affinity Modification of the Restriction Endonuclease SsoII by 2′-Aldehyde-Containing Double Stranded DNAs

“…This approach employs DNA ligands carrying a reactive group in 2′ position of the sugar moiety. Recent studies in our laboratory on a series of substrate analogs containing an aldehyde group in the 2′ position of the sugar moiety revealed their potential applicability for affinity modification of p50 subunit of human transcription factor NF κB [9,14].…”

“…DNA duplexes I V were prepared as described in [9]. The DNA duplexes I, II, and V contained 5′ 32 P label in both strands, whereas duplexes III and IV contained the label in the modified strand.…”

Use of Carbonyl Group Addition−Elimination Reactions for Synthesis of Nucleic Acid Conjugates

“…The duplex structure was somewhere between A-and B-forms, as judged by CDspectroscopy (65,68). More recently, oligonucleotides containing an aldehyde group at the 2′-position of a sugar residue were used for affinity modification of NF-kB protein (69,70) and DNA-methyltransferase SsoII (Vorobjeva, Zatsepin, Kubareva, Oretskaya, unpublished results). Also oligonucleotides with 2′-dialdehyde group were used in affinity modification of methyltransferases ( 71) and T7 RNA polymerase (72).…”