DNA structures

As the present book is concerned with the free-radical chemistry of DNA, it is worth mentioning that hydroxyl radical footprinting, based on the forma- 

3 Analysis with a fluorescence microscope of different parameters that characterise the DNA structure. 

DNA is composed of three chemical functions A deoxyribose (a pentose, i.e., a sugar with five carbons), organic (nitrogenous) bases (pyrimidines cytosine and thymine purines adenine and guanine), and a phosphoric acid. 

Ribose and deoxyribose contribute to the formation of RNA and DNA, respectively. Binding of a base to the pentose yields a nucleoside, and binding of a phosphoric acid to the nucleoside forms a nucleotide. Finally, different nucleotides bound together form a nucleic acid. There is a specific complementary nature to the bases adenine with thymine and guanine with cytosine. The C-G pair has three hydrogen bonds, while the A-T pair has only two, thus preventing incorrect pairing. The sequence of the bases determines the primary structure of the DNA. 

An alternative conformation of the B DNA appears in solution when the amount of water necessary to hydrate the double helix is not sufficient. In the A conformation, the pitch is 24.6 A, and a complete turn of the helix needs the presence of 11 base pairs. Finally, we should add that in vivo, we do not know whether the A form of DNA does really exist. The third conformation of DNA is the Z form where the helix is twisted toward the left. 

The sequence of DNA bases is of great importance in heredity. In fact, the sequence of bases in one strand has a complementary relationship to the sequence of bases in the other strand. In other terms, information contained in the sequence of one strand is conserved in the second strand. 

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Hippel, P.H. von, Wang, K.-Y. Dynamic aspects of native DNA structure kinetics of the formaldehyde reaction with calf thymus DNA. J. Mol. Biol. 61 (1971) 587-613... 

Packer M J, M P Dauncey and C A Hunter 2000. Sequence-dependent DNA Structure Dinucleotide Conformational Maps. Journal of Molecular Biology 295 71-83. 

The first six chapters of this book deal with the basic principles of protein structure as we understand them today, and examples of the different major classes of protein structures are presented. Chapter 7 contains a brief discussion on DNA structures with emphasis on recognition by proteins of specific nucleotide sequences. The remaining chapters illustrate how during evolution different structural solutions have been selected to fulfill particular functions. 

Early diffraction photographs of such DNA fibers taken by Rosalind Franklin and Maurice Wilkins in London and interpreted by James Watson and Francis Crick in Cambridge revealed two types of DNA structures A-DNA and B-DNA. The B-DNA form is obtained when DNA is fully hydrated as it is in vivo. A-DNA is obtained under dehydrated nonphysiological conditions. Improvements in the methods for the chemical synthesis of DNA have recently made it possible to study crystals of short DNA molecules of any selected sequence. These studies have essentially confirmed the refined fiber diffraction models for A- and B-DNA and in addition have given details of small structural variations for different DNA sequences. Furthermore, a new structural form of DNA, called Z-DNA, has been discovered. 

Figure 8.23 The helix-turn-helix motifs of the subunits of both the PurR and the lac repressor subunits bind to the major groove of DNA with the N-terminus of the second helix, the recognition helix, pointing into the groove. The two hinge helices of each arm of the V-shaped tetramer bind adjacent to each other in the minor groove of DNA, which is wide and shallow due to distortion of the B-DNA structure. (Adapted from M.A. Schumacher et al.. Science 266 763-770, 1994.)...

The central 10 base pairs of the palindromic DNA molecule have a regular B-DNA structure. Between base pairs 5 and 6 in each half of the fragment (base pairs are counted from the center) there is a 40° kink which causes these base pairs to be unstacked (Figure 8.24a). After this localized kink the two end regions have an essentially B-DNA structure. The kink occurs at a TG step in the sequence GTG. These TG steps at positions 5 and 6 are highly conserved in both halves of different CAP-binding sites, presumably in part because they facilitate kinking. 

In the two complexes studied by x-ray crystallography, the interactions between TBP and the DNA, as well as the deformation of the B-DNA structure, are very similar, and we will illustrate some of these details for the yeast structure. Minor details of the two complexes vary due to differences in some of the side chains and nucleotides that are present in the interaction areas. 

In conclusion, one important factor that contributes to the strong affinity of TBP proteins to TATA boxes is the large hydrophobic interaction area between them. Major distortions of the B-DNA structure cause the DNA to present a wide and shallow minor groove surface that is sterically complementary to the underside of the saddle structure of the TBP protein. The complementarity of these surfaces, and in addition the six specific hydrogen bonds between four side chains from TBP and four hydrogen bond acceptors from bases in the minor groove, are the main factors responsible for causing TBP to bind to TATA boxes 100,000-fold more readily than to a random DNA sequence. 

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. 

TFIIA and TFIIB are two basal transcription factors that are involved in the nucleation stages of the preinitiation complex by binding to the TBP-TATA box complex. Crystal structures of the ternary complex TFIIA-TBP-TATA box have been determined by the groups of Paul Sigler, Yale University, and Timothy Richmond, ETH, Zurich, and that of the TFIIB-TBP-TATA box by Stephen Burley and collaborators. The TBP-DNA interactions and the distortions of the DNA structure are essentially the same in these ternary complexes as in the binary TBP-TATA complex. 

Interactions that are not sequence specific are also an Important part of the binding and occur between the sugar and phosphate residues of the DNA and the side-chain and main-chain atoms of the protein. In the crystals the DNA fragment retains the B-DNA structure with only minor distortions. 

Thirty percent of the tumor-derived mutations are in L3, which contains the single most frequently mutated residue, Arg 248. Clearly the interaction between DNA and the specific side chain of an arginine residue inside the minor groove is of crucial importance for the proper function of p53. It is an open question whether this interaction is needed for the recognition of specific DNA sequences, or is required for the proper distortion of the DNA structure, or a combination of both. Other residues that are frequently mutated in this region participate in interactions with loop L2 and stabilize the structures of loops L2 and L3. Mutations of these residues presumably destabilize the structure so that efficient DNA binding can no longer take place. 

The helical basic region of Max binds to the major groove of DNA in the same manner as MyoD (see Figures 10.26 and 10.28). The DNA structure is essentially that of B-DNA with minor distortions as in the MyoD complex. Both Max and MyoD recognize the same consensus half-site DNA sequence 5 -CAC-3 ... 

FIGURE 12.14 Comparison of the deoxy-guanosine conformation in B- and Z-DNA. In B-DNA, the Cl -N-9 glycosyl bond is always in the anti position (lefi). In contrast, in the left-handed Z-DNA structure, this bond rotates (as shown) to adopt the syn conformation. 

FIGURE 12.16 The structures of ethidiutn bromide, acridine orange, and actinomycin D, three intercalating agents, and their effects on DNA structure. 

Thomson Click Organic Interactive to use interactive animations to view aspects of DNA structure. 

The DNA structure involves two polyanionic phosphodiester strands linked together by hydrogen bonding of base pairs. The strands can be separated by a denaturation process (melting). The melting temperatnre increases with an increase in guanine (G)-cytosine (C) content, since this base pair possess three hydrogen bonds as compared to just two for the adenine (A)-thymine (T) pair. 

We will briefly discuss the properties of the structures obtained as a result of DNA/polymer interactions, with emphasis on electrostatic ones. The reason for this preference is purely practical, since it has been a major route for the preparation of various DNA structures involved in gene transfer in vitro and, more recently, in vivo. 

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