Proteins DNA interactions with

Brockman JM, Fmtos AG, Com RM (1999) A multistep chemical modification procedure to create DNA arrays on gold surfaces for the study of protein-DNA interactions with surface plasmon resonance imaging. J Am Chem Soc 121 8044-8051... 

Guidelines for Studying Protein-DNA Interactions with the BIAcore... 

To study a novel protein-DNA interaction with the BIAcore an initial characterization of the interaction by a gel-shift and/or footprint assay will be required [2,3]. These assays provide data on the stoichiometry of the protein-DNA complex, the specificity of the interaction, and the localization of the binding site on the DNA. 

The specific protein-DNA interactions described in this book are all with DNA in its regular B-form, or, in some cases with distorted B-DNA. In biological systems DNA appears not to adopt the A conformation, although double-stranded RNA does preferentially adopt this conformation in vivo. Whether or not Z-DNA occurs in nature is a matter of controversy. However, the formation of A-DNA and Z-DNA in vitro does illustrate the large structural changes that DNA can be forced to undergo. 

The x-ray structure of DNA complexes with 434 Cro and repressor revealed novel features of protein-DNA interactions... 

The protein-DNA interactions have been analyzed in detail at high resolution in the complex between the 434 repressor fragment and the ORl containing 20mer DNA. A pseudo-twofold symmetry axis relates the halves of this complex. The symmetry is not exact since the nucleotide sequence of the DNA is slightly different in each half (see Table 8.2). However, the interactions between one protein subunit and one half of the DNA are very similar to those between the second subunit and the other half of the DNA since most of the bases that interact with the protein are identical in both halves. Details of the interaction are very similar to those in the complex with the palindromic synthetic 14mer of DNA shown in Figures 8.14 and 8.15. The base pairs at one end of the DNA, 1-14, 2-13, etc. are called base pairs 1, 2, etc. 

Figure 8.15 Sequence-specific protein-DNA interactions provide a general recognition signal for operator regions in 434 bacteriophage, (a) In this complex between 434 repressor fragment and a synthetic DNA there are two glutamine residues (28 and 29) at the beginning of the recognition helix in the helix-turn-helix motif that provide such interactions with the first three base pairs of the operator region.

The sharp bend of DNA at the TATA box induced by TBP binding is favorable for the formation of the complete DNA control module in particular, for the interaction of specific transcription factors with TFIID. Since these factors may bind to DNA several hundred base pairs away from the TATA box, and at the same time may interact with TBP through one or several TAFs, there must be several protein-DNA interactions within this module that distort the regular B-DNA structure (see Figure 9.2). The DNA bend caused by the binding of TBP to the TATA box is one important step to bring activators near to the site of action of RNA polymerase. 

The NMR study by Wiithrich and coworkers has shown that there is a cavity between the protein and the DNA in the major groove of the Antennapedia complex. There are several water molecules in this cavity with a residence time with respect to exchange with bulk water in the millisecond to nanosecond range. These observations indicate that at least some of the specific protein-DNA interactions are short-lived and mediated by water molecules. In particular, the interactions between DNA and the highly conserved Gin 50 and the invariant Asn 51 are best rationalized as a fluctuating network of weak-bonding interactions involving interfacial hydration water molecules. 

The GAL4 recognition module therefore contains only one protein side chain, Lys 18, that provides specific interactions with the DNA. The remaining specific interactions with DNA are from main-chain atoms and depend critically on the correct conformation of the protein. The correct positioning of the C-terminus of the a helix is particularly important for recognition. This is to date the only example of a protein-DNA interaction in which... 

Instead of activating transcription the cortisol-induced GR represses IL-6 synthesis and, even more surprisingly, repression does not involve the GRE elements, but rather the kB site (Fig. 1). It appeals that of a monomeric GR protein without itself touching the DNA interacts with the RelA component of NF-kB [3]. As a result GR blocks the action of NF-kB. The negative interference by this crosstalk is not restricted to NF-kB, it occurs also with AP-1 and CREB, and with several other transcription factors not relevant for IL-6 expression. A nuclear isoform of the LIM protein Trip6 mediates the interaction between these factors and is required for the inhibitory GR function. This interesting negative crosstalk is part of the immune-suppressive action of cortisol. 

Gebara, M. M., and McCarrey, J. R. (1992). Protein-DNA interactions associated with the onset of testis-specific expression of the mammalian Pgk-2 gene. Mol. Cell. Biol. 12 1422-1431. 

Solomon MJ, Larsen PL, Varshavsky A. Mapping protein-DNA interactions in vivo with formaldehyde evidence that histone H4 is retained on a highly transcribed gene. Cell 1988 17 937-947. 

Fig. 1.6. Binding domain to DNA. ERs contain two structures called zinc fingers, typical of proteins that interact with DNA. One zinc atom forms four links of coordination with four cysteine residues of the protein structure, which occupy nearby positions, thus leaving a loop of some 15 to 22 aminoacids. The zinc fingers of the receptor are capable of interacting with specific sequences of DNA, the hormone response elements, with which they establish hydrogen bridges and form stable structures...

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