DNA-binding protein

Another class of DNA-binding proteins are the polymerases. These have a nonspecific interaction with DNA because the same protein acts on all DNA sequences. DNA polymerase performs the dual function of DNA repHcation, in which nucleotides are added to a growing strand of DNA, and acts as a nuclease to remove mismatched nucleotides. The domain that performs the nuclease activity has an a/P-stmcture, a deep cleft that can accommodate double-stranded DNA, and a positively charged surface complementary to the phosphate groups of DNA. The smaller domain contains the exonuclease active site at a smaller cleft on the surface which can accommodate a single nucleotide. 

Figure 8.3 The DNA-binding protein Cro from bacteriophage lambda contains 66 amino acid residues that fold into three a helices and three P strands, (a) A plot of the Ca positions of the first 62 residues of the polypeptide chain. The four C-terminal residues are not visible in the electron density map. (b) A schematic diagram of the subunit structure. a helices 2 and 3 that form the helix-turn-helix motif ate colored blue and red, respectively. The view is different from that in (a), [(a) Adapted from W.F. Anderson et al., Nature 290 754-758, 1981. (b) Adapted from D. Ohlendorf et al., /. Mol. Biol. 169 757-769, 1983.]...

However, the a helices are not packed against each other in the usual way as described in Chapter 3. Instead, a helices 2 and 3, residues 15-36, form a unique helix-turn-helix arrangement that in 1981 had only been observed once, in a different bacterial DNA-binding protein, the catabolite gene-activating protein CAR... 

The presence of this common helix-turn-helix motif poised for DNA binding in lambda Cro and repressor provided considerable stimulus for further genetic and structural studies of these and other procaryotic DNA-binding proteins. All the results essentially supported the proposed mode of binding between these regulator proteins and DNA. 

Figure 8.17 Schematic diagram of the main features of the interactions between DNA and the helix-turn-helix motif in DNA-binding proteins.

Some of the procaryotic DNA-binding proteins are activated by the binding of an allosteric effector molecule. This event changes the conformation of the dimeric protein, causing the helix-tum-helix motifs to move so that they are 34 A apart and able to bind to the major groove. The dimeric repressor for purine biosynthesis, PurR, induces a sharp bend in DNA upon binding caused by insertion of a helices in the minor groove between the two... 

Sauer, R.T., et al. Homology among DNA-binding proteins suggests use of a conserved super-secondary structure. Nature 298 447-451, 1982. 

The homeodomain frequently binds to DNA as a monomer, in contrast to procaryotic DNA-binding proteins containing tbe belix-turn-helix motif, which usually bind as dimers. In vitro tbe homeodomain binds specifically to... 

Figure 9.8 Schematic diagram of the three-dimensional structure of the Antennapedia homeodomain. The structure is built up from three a helices connected by short loops. Helices 2 and 3 form a helix-turn-hellx motif (blue and red) similar to those in procaryotic DNA-binding proteins. (Adapted from Y.Q. Qian et al.. Cell 59 573-580, 1989.)...

Phillips, S.E.V. Built by association structure and function of helix-loop-helix DNA-binding proteins. Strueture 2 1-4, 1994. 

The world of zinc-containing DNA-binding proteins is by no means exhausted by these three subfamilies. Several other subfamilies are already known with different three-dimensional structures and different sequence patterns of cysteine and histidine residues that form the zinc ligands. Further subfamilies may well be discovered as the genomes of different species are sequenced whether or not any fundamentally new principles for DNA-protein recognition will be discovered amongst these new subfamilies remains to be seen. 

The leucine zipper DNA-binding proteins, described in Chapter 10, are examples of globular proteins that use coiled coils to form both homo- and heterodimers. A variety of fibrous proteins also have heptad repeats in their sequences and use coiled coils to form oligomers, mainly dimers and trimers. Among these are myosin, fibrinogen, actin cross-linking proteins such as spectrin and dystrophin as well as the intermediate filament proteins keratin, vimentin, desmin, and neurofilament proteins. 

Histone H3 Histones are DNA-binding proteins found in chromosomes 135 amino acid residues. Note die very basic nature of this protein dne to its abmidance of Arg and Lys residues. It also lacks tryptophan. 

It is now apparent that bacteria have developed resistance to heavy metals and the detoxifying process is initiated and controlled by metallo-regulatory proteins which are able selectively to recognize metal ions. MerR is a small DNA-binding protein which displays a remarkable sensitivity to Hg +. The metal apparently binds to S atoms of cysteine and this has been a major incentive to recent work on Hg-S chemistry. 

Originally discovered as DNA-binding proteins that mediate interferon signaling, recent data demonstrated that STAT1 can also exert constitutive functions in the nucleus, which do not require STAT activation with tyrosine phosphorylation. Cells lacking STAT1 are... 

A leucine zipper is a structural motif present in a large class of transcription factors. These dimeric proteins contain two extended alpha helices that grip the DNA molecule much like a pair of scissors at adjacent major grooves. The coiled-coil dimerization domain contains precisely spaced leucine residues which are required for the interaction of the two monomers. Some DNA-binding proteins with this general motif contain other hydrophobic amino acids in these positions hence, this structural motif is generally called a basic zipper. 

Nuclear factor kappa B (NF-kB) is the generic term for a family of dimeric eukaryotic transcription factors, composed of members of the Rel family of DNA-binding proteins including the mammalian proteins RelA (or p65), cRel, RelB, p50 and p52, and the Drosophila proteins Dorsal, Dif and Relish. These proteins bind with different affinities to a consensus DNA sequence motif (called the kB site) consisting of the sequence 5 -GGGRNNYYCC-3 in which R is a purine, Y is a pyrimidine, and N is any base. 

The core unit of the chromatin, the nucleosome, consists of histones arranged as an octamer consisting of a (H3/ H4)2-tetramer complexed with two histone H2A/H2B dimers. Accessibility to DNA-binding proteins (for replication, repair, or transcription) is achieved by posttranslational modifications of the amino-termini of the histones, the histone tails phosphorylation, acetylation, methylation, ubiquitination, and sumoyla-tion. Especially acetylation of histone tails has been linked to transcriptional activation, leading to weakened interaction of the core complexes with DNA and subsequently to decondensation of chromatin. In contrast, deacetylation leads to transcriptional repression. As mentioned above, transcriptional coactivators either possess HAT activity or recruit HATs. HDACs in turn act as corepressors. 

Sorger, P.K. Pelham, H.B.R. (1988). Yeast heat shock factor is an essential DNA binding protein that exhibits temperature-dependent phosphorylation. Cell 54, 855-864. 

Tab. 3.1 DNA-binding proteins that have been inhibited by polyamides. The known DNA binding motifs from NMR or crystal structure data are shown. Significant groove contacts and proposed mechanism of polyamide inhibition are also shown for each protein...

---