Meaning Of Embovang

Note that the low displacement and tilt of B-DNA indicates that the bases sit on the axis and are perpendicular to it (Figure 16.5).


Figure 16.5: Structure of the A-DNA, B-DNA and Z-DNA forms of the DNA double helix. The B-form is thought to be the most prevalent in vivo. M = major groove, m = minor groove, G = single groove in Z-DNA. (Modified from DNA Replication. Kornberg and Baker (1992), WH Freeman, New York.)


Figure 16.5: Structure of the A-DNA, B-DNA and Z-DNA forms of the DNA double helix. The B-form is thought to be the most prevalent in vivo. M = major groove, m = minor groove, G = single groove in Z-DNA. (Modified from DNA Replication. Kornberg and Baker (1992), WH Freeman, New York.)

Local flexibility in DNA structure. The analysis of oligonucleotide crystals as opposed to fibres shows that there is great variation in the helical parameters of molecules with diverse base sequences. This occurs because different base sequences influence helical and torsional parameters to maximize the stability of stacking and pairing interactions. B-DNA is particularly flexible in this respect and different local conformations form to adapt to particular sequences. This indicates that DNA probably does not exist in rigid conformational forms but may change smoothly between different conformations punctuated by local polymorphisms such as bent DNA and helical transitions (sudden transitions between different helical conformations within a single molecule, e.g. B-Z transitions). DNA bending is an intrinsic property depending on stacking interactions which, according to local sequence, may be isotropic (unbiased) or anisotropic (bending in a specific direction), Intrinsic DNA bends occur in AT-rich runs and in repeats of the sequence GGCC in step with helical periodicity. DNA bending can also be induced by proteins (see Nucleic Acid-Binding Proteins) and by circularization {q.v. DNA topology). Induced bending is necessary for DNA packaging in chromosomes (see Chromatin) and for replication, recombination and transcription (q.v.). Proteins may also recognise DNA that is bent in a certain way (e.g. top ois o m era ses).

Secondary structure in RNA and nonduplex DNA. In RNA and single-stranded regions of DNA, secondary structure is determined by intramolecular base pairing. Since cellular DNA is usually present as a duplex, the bases are available for intramolecular interactions only rarely. Conversely, intramolecular secondary structures are abundant in cellular RNA and underlie their functional specialization. RNA secondary structures play a major role in gene expression and its regulation: base pairing between rRNA and mRNA controls the initiation of protein synthesis, base pairing between tRNA and mRNA facilitates translation, RNA hairpins and stem loops control transcriptional termination, translation efficiency and mRNA stability, and RNA-RNA base pairing also plays a major role in the splicing of introns (see Transcription, RNA Processing, Protein Synthesis). The major classes of intramolecular nucleic acid secondary structures are listed in Table 16.4. Like DNA, RNA helical conformation is modulated by local sequence character, but the relatively high percentage of modified bases further adds to the variety of structures which form.

Table 16.4: Intrastrand nucleic acid secondary structure elements

Secondary structure


Bulges and bulge loops

Internal loops (bubbles)

Hairpins, stem loops

Panhandle Cruciform

Deformities on one side of a duplex which has excess residues. A bulge is caused by a single excess residue, and bulge loops by more than one, These distort stacking of neighboring bases and induce a bend, increasing the accessibility of the major groove. Bulges and bulge loops in double-stranded DNA are caused by insertions (see Mutagenesis and DNA Repair) Deformities caused by one or more mismatching base pairs in an otherwise duplex structure. In RNA, internal loops or bubbles have been Implicated as protein recognition sites Secondary structures which may form in regions of hyphenated dyad symmetry. The two complementary regions base pair to form a stem which may fold over on itself (hairpin) or end in a loop of unpaired nucleotides (stem-loop, hairpin-loop). Three and four nucleotide loops are particularly stable structures due to special base pairing and stacking interactions within the loop. Hairpins and stem loops are key functional elements in many biological systems, including, for example, tRNA and rRNA structure, transcriptional termination, control of translation and packaging of viral genomes, suggesting that they are sites for protein-RNA Interaction A discrete linear nucleic acid whose termini are complementary and form a short duplex region, the rest of the molecule forming a loop In double-stranded nucleic acids with regions of dyad symmetry, a cross-shaped structure which forms when hairpins or stem loops arise simultaneously in both strands. An important source of replication errors, such structures are repressed by single-strand binding proteins

Lariats (q.v.) are often classed as secondary structures but, because they are formed by the covalent bonds joining nucleotides, they are strictly primary structures.

16.3 Nucleic acid tertiary structure

Tertiary strand interactions in DNA. Nucleic acid tertiary structures reflect interactions which contribute to overall three-dimensional shape. This includes interactions between different secondary structure elements, interactions between single strands and secondary structure elements, and topological properties of nucleic acids.

In DNA, tertiary interactions involve single strands interacting with duplexes or duplexes interacting with duplexes, resulting in the formation of triple and quadruple strand structures. Guanine can form base tetrads, and DNA containing runs of guanosine residues can form quadruplex structures which may contribute to telomere structure (q.v.). Triple-stranded DNA forms spontaneously when a single strand interacts with bases in a duplex molecule; the interactions occur through the major groove and involve the formation of nonWatson-Crick base pairs between the invading strand and one of the resident strands. Studies of the interactions of oligonucleotides with duplex DNA provided the first evidence for triple helices, and identified four common types of base triples where Hoogsteen base pairing is involved. H-DNA is a form of intramolecular triple-stranded DNA (so-called because it is protonated) which arises in paired homopurine/homopyrimidine sequences and involves Hoogsteen base pairs. The physiological role of H-DNA is unclear, although it is implicated in the regulation of some genes, e.g. GAP-43 in mammals (also q.v. triple-helix therapy). Triple-stranded DNA also forms during recombination when a single strand invades a duplex. Because of topological constraints (see next section) an intact invading strand must pair with the complementary strand in the duplex without winding around it. Such a structure is a paranemic joint, and must be stabilized by proteins. It may involve extensive unwinding of the target duplex, or the formation of alternative segments of left- and right-handed helix (V-DNA). If the invading strand has a free end, or further topoisomerase activity, the invading strand winds around its complementary partner in the duplex in the norma! fashion to form a plectonemic joint — the resident strand is ejected as a displacement loop (D-loop). Similar tertiary structures termed R-loops form when RNA transcribed from duplex DNA is stabilized in situ, as occurs, for example, during the priming of replication in the ColEl plasmid (see Plasmids). Four-strand tertiary structures, Holliday junctions, also form during recombination (q.v. homologous recombination).

Tertiary strand interactions in RNA. RNA folds into complex structures involving tertiary interactions between strands, loops, and duplexes. For example, in tRNA there are examples of base triples, sections of triple helix, stem junctions (where two or more duplex regions are joined) and pseudo-knots (where strands interact with stem-loops).

RNA folding is often controlled by molecular chaperones (q.v.) like protein folding. The complexity of RNA tertiary structure allows it to form biologically active molecules, and like proteins, RNA can catalyze biochemical reactions. Such catalystic RNAs are termed ribozymes. Some ribozymes are autocatalytic (e.g. the transcripts of self-splicing introns, q.v,). Others are trans-acting, including ribonuclease P and the family of hammerhead ribozymes found in some plant viroids (q v,), so called because of the three-helical structure of the catalytic domain (also q.v. gene therapy).

DNA topology. Topology is the branch of mathematics dealing with the properties of geometric structures which are independent of size and shape and unchanged by deformation. If a double-stranded DNA molecule has free ends (e.g. a linear molecule), the two strands wind around each other in the most energetically favorable manner, and the molecule is said to be relaxed. The number of times one strand winds around the other in this relaxed state is the duplex winding number. If extra twists are introduced into such a molecule to make it overwound, then the total number of helical turns — which is the linking number — exceeds the duplex winding number. Conversely, if twists are removed from the molecule to make it underwound, the duplex winding number exceeds the linking number. In either case, the strands can rotate with respect to each other and return the molecule to its relaxed state. In a closed circle, however, there are no free ends and the linking number is a topological property — it can be changed only by breaking the circle open, not by deforming it. If DNA in a closed circle becomes overwound or underwound, the only way to relax the torsional strain thus produced is by supercoiling, where a twist is introduced into the helical axis itself. Supercoiling is another form of nucleic acid tertiary structure, one involving the effect of torsional stress upon shape rather than strand-strand interactions (Box 16.2).

The physiological significance of supercoiling is that unconstrained DNA is often biologically inactive. Negative supercoiling is required for many essential processes: replication, transcription and recombination included. Supercoiled DNA has stored energy which drives these reactions. In eukaryotes, which possess linear chromosomes, topological constraints are introduced by organizing chromatin into loops with ends fixed by scaffold proteins; nucleosomes introduce negative supercoils into eukaryote DNA (q.v. chromatin loops, matrix associated region, DNA topoisomerase, site-specific recombination).

Nucleic acid quaternary structure. In many structures, nucleic acids interact in trails (e.g. the ribo-some and spliceosome), and this may be considered a quaternary level of nucleic acid structure. Nucleic acids also Interact with an enormous number of proteins (e.g. genome structural proteins, transcription factors, enzymes, splicing factors). Many of these proteins have a significant effect on DNA or RNA conformation. Interactions with proteins may be general or sequence specific, and may involve subtle or overt changes in structure. The restriction endonucleases EcoRI and EcoRV, for instance, both introduce a pronounced kink in the DNA at their recognition sequence which may facilitate their endonucleolytic activity. Proteins of the HMG class appear specifically to bend DNA in order to facilitate interactions between components bound at distant sites. For further discussion of nucleic acid-protein interactions see Nucleic Acid-Binding Proteins.

Box 16.1 : Helix morphology, parameters and torsion angles

Helix morphology, DNA and RNA helices are classified according to their gross morphology, Criteria include helical diameter, helical sense (direction of rotation}, pitch (number of base pairs per helical turn) and the width and depth of the major and minor grooves. The grooves were originally defined on the basis of their relative sizes in B-DNA. However, because both can change size under conformational stress, a precise definition is needed. The major groove is defined as that containing the C4 of a pyrimidine or N7 of purine, whereas the minor groove contains the O2 of a pyrimidine or the N3 of a purine.

Torsion angles. Details of conformational structure can be described unambiguously in terms of the torsion angles of the bonds in the sugar-phosphate backbone, in the furanose ring itself, and of the glycoside bond, as shown in the figure below. Many of these angles are interdependent and conformational descriptions can be abbreviated to the specification of five bonds: 6 (C^-C4', which is related to sugar pucker), % (the glycosidic bond), y (C4'-C5') and the phosphoester bonds a and £,. Additionally, these bonds can be described in terms of gross conformation rather than absolute angle, e.g. by describing the conformation of the glycosidic bond as anfr or syn and by describing the sugar pucker as C3' endo or C2' endo.

Helical parameters. Details of the positional relationship between stacked and paired bases are described in the terms of a universal set of helix parameters. There are translational (displacement) and rotational parameters which describe the relative position of bases in base pairs, the relative position of successive base pairs in base stacks, and the absolute position of base pairs relative to the helical axis. The terms used are given below.


Coordinates x-axis y-axis

Absolute position relative to helical axis x-displacement y-displacement

Displacement of successive base pairs Shift Slide

Displacement of baes within pairs Shear Stretch

Absolute rotation relative to helical axis Inclination (it) Tip (0)

Rotation of successive base pairs Tilt (t) Roll (p)

Rotation of bases within pairs Buckle (kJ Propeller twist (a>)

Rise Stagger

Box 16.2: Quantifying the topological properties of DNA

Measuring helical winding. The linking number

(i.) is the number of times one DNA strand wraps round the other in a duplex, and for right-handed helices, L is positive. The duplex winding number (Lq) is the linking number for relaxed DNA and represents the most energetically favorable configuration. For B-DNA, the average L0 = n/10.3 where n is the number of base pairs. If DNA is relaxed DNA, L = L0, but any deviation from this state by overwinding or underwinding creates torsional strain. In open DNA (DNA with free ends), the strain is countered by rotation of the strands relative to each other, whereas in covalently closed DNA (circular DNA or DNA with fixed ends) oppositional rotation is prevented and torsional strain must be countered by supercoiling.

Measuring supercoiling. The degree of super-coiling in a given DNA molecule is expressed as the superhelical density [X), which is calculated as follows:

"Rie superhelix winding number (t) is the difference between L and Lq. If DNA is overwound, positive supercoils are introduced and t is positive, whereas underwound DNA generates negative supercoils

1 and 11s negative, t quantifies the degree of torsional strain a given molecule is under and thus its propensity to undergo supercoiling, but it does not measure the actual number of superhellcal turns, because the pitch of the helix may also be changed by torsional strain. The number of superhelical turns is expressed as the writhing number (IV). This is related to the linking number in the equation L = T + W where T, the twisting number, is the total number of turns in a DNA molecule. The linking number is topological (i.e. invariable under deformation) so any change in IV, the number of turns of superhelix, must be countered by an equal and opposite change In T, In a relaxed molecule, L = T, hence IV = 0 and all turns are helical turns. One unit of writhe is equivalent to one half superhelical turn, i.e. a turn l of 180° in the helical axis of the DNA. Each unit of writhe can be thought of as a point at which two ' duplexes cross each other when a supercoiled molecule is forced to lie on a flat surface, such a point being described as a node.

Catenation and knotting. As well as helical winding within a closed molecule, catenation (the Interlocking of DNA circles) and the formation of knots are also topological properties of DNA. In neither case can such structures be resolved without breaking the DNA molecule open to untangle it, and both structures involve nodes where duplexes must cross each other when the molecules are placed flat. The total amount of linking in a given DNA molecule Is thus expressed as its linking number L plus the amount of knotting and catenation, i.e. total linking = J. + C + K. j


Blackburn, G.M, and Gait, M.J. (eds) (1996) Nucleic and function. In: DNA Replication. 2nd edn, pp.

Acids in Chemistry and Biology. Oxford University 1-52. W.H. Freeman, New York.

Press, Oxford. Rich, A. el al. (1984) The chemistry and biology of left-

Dickerson, R.E, el al. (1989) Definitions and «omenda- handed Z-DNA. Annu. Rev. Biochem. 53: 791-846.

ture of nucleic acid structure parameters. EMBO /, Vang, Y„ Westcott, T.P., Pedersen, S.C., Tobias, I. and

8:1-4. Olson, W.K. (1995) Effects of localised bending on Romberg, A. and Baker, T.A. (1992) DNA structure DNA supercoiling. Trends Biochem. Sci. 20: 313-319.

Further reading

Doudna, J.A. and Cate, J.M. (1997) RNA structure: crystal clear? Curr. Op. Struct, Biol. 7: 310-316.

Eaton, B.E. and Pieken, W.A. (1995) Ribonucleosides and RNA. Annu. Rev. Biochem. 64: 837-863.

Frank-Kamenetskii, M.D. and Mirkin, S.M. (1995) Triplex DNA structures. Rev. Biochem. 65:

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