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Some Structural Aspects of Non-covalent DNA Binding Agents


Keywords: Drug-DNA binding, DNA intercalation, Minor Groove binding, Drug Design
Introduction
Cancer is a group of over 100 diseases known by the prolific increase of cellular mass of the tissues in an unusual manner to cause death of a person.1 In addition, cancerous cells do not adhere to each other and to the inter-cellular matrix. This property of non-adherence enables them to propagate to distant sites in the body and spread the cancer.2
            Modern day drug discovery is fundamentally driven by biology. Biological molecules like DNA & proteins are valuable targets for drug action. DNA is central to several life processes like replication, transcription, etc. DNA produces copies of itself via replication mechanism. On the other hand, DNA transcription results in the synthesis of proteins. Since DNA is an indispensable part of the living system, it can be a valuable target for drug action.3 With the help of DNA binding drugs, one can target vital processes like replication, protein synthesis and cell division which ultimately lead to the treatment of cancer.
Structure of DNA
In order to target DNA, it is essential to understand the structural features of DNA. DNA is a very long thread like polymer composed of a linear array of monomers called nucleotides. The critical feature of DNA is its linear order of the four nucleotides, which are the storehouse of all the encoded information. Each nucleotide is composed of an aromatic base (a purine or pyrimidine ring), deoxyribose sugar and a phosphate group (Fig 1). The hydrogen-bonding surface of the individual bases is inside, at the center of the double helix. The phosphate backbone is outside the double helix.

Figure 1: A nucleotide of DNA showing nitrogenous base, pentose sugar and a phosphate group.
            There are three major forms of DNA commonly found in nature, viz. A, B and Z forms. Main structural features of A, B and Z forms are shown in table 1. B-form DNA is the most common structure type found in humans.
Table 1: Structural Features of A, B and Z DNA double helix

B DNA
Z DNA
A DNA
Helix sense
Right-handed
Left handed
Right handed
Rotation/bp
35.9°
60°/2
33.6°
Mean bp/turn
10.0
12
10.7
Inclination of bp to axis
-1.2°
-9°
+19°
Rise/bp along axis
3.32Å
3.8 Å
2.3 Å
Pitch/turn of helix
33.2Å
45.6 Å
24.6 Å
Mean propeller twist
+16°
+18°
Glycosyl angle
anti
C: anti, G: syn
anti
Sugar pucker
C2'-endo
C: C2'-endo,
G: C2'-exo
C3'-endo
Diameter
20 Å
18 Å
26 Å

Mode of Action of Non-covalent DNA binding drugs
There are two principal modes of drug-DNA binding, viz. Intercalation and minor groove binding. Majority of drug molecules interact with DNA by either of these two methods. Exceptions, however, include drugs which bind in the major groove and to the sugar-phosphate backbone of DNA.
 Intercalative binding
Intercalation can be described as the insertion of drug molecule or a part of it between the base pairs of the DNA double helix (Fig 2).

Figure 2: Drug (magenta) showing intercalation between the base pairs of DNA.
            Bases in each DNA strand are stacked over each other via pi-pi stacking forces. Since the DNA double helix is stable in its structure and configuration, inserting any foreign molecule between the base pairs would disturb its original shape. The incorporation of drug molecule causes unstacking of base pairs leading to the opening of base pairs along with the unwinding of strands of DNA. In general, it is observed that the base pairs next to the intercalation site are not occupied by the drug molecule due to the lack of intercalation cavity in these base pairs.
            The main structural feature for the intercalating drugs is the presence of planar ring systems. Normally, a compound containing two or more six membered fused aromatic rings is capable of intercalation between the DNA base pairs. Stacking forces are essential for a stable drug-DNA intercalation complex. Fused aromatic rings possess enough stacking ability to bind with DNA bases. In addition, the compound may also possess a positively charged atom (like N+) to form cation-pi interactions to facilitate binding. Compounds like Ellipticine4 and Cryptolepine5 are known DNA intercalators.
 Minor Groove binding
Apart from the inward facing base pairs, DNA structure also possesses two groves (major and minor grooves) which run though the entire length of the macromolecule (Fig 3). These grooves differ significantly in their electrostatic potentials, steric effects and hydration characteristics. Structurally, minor groove is narrower and deeper than the major groove.

Figure 3: DNA molecule showing inward facing base pairs and major & minor grooves.
            Major groove possesses greater area and volume than the minor groove. This causes small drug like molecules to slip-off from the major groove side of DNA. The minor groove, however, due to its shape provides enough motifs for small drug like molecules to form hydrogen bonding and van-der-Waal’s contacts with the atoms of DNA to form a comlex.
            Drugs which bind in the minor groove possess a crescent shaped structure which fits according the shape of the minor groove. Typically, minor groove binding molecules have several simple aromatic rings connected by bonds with torsional freedom. This facilitates compounds with the appropriate twist, to fit into the helical curve of the minor groove. Such minor groove binding is accomplished by the displacement of water from the groove and forming van der Waal’s contacts with the helical atoms present in the wall of the minor groove.
            Additional specificity in the binding is derived from contacts between the bound molecule and the edges of the base pairs on the ‘floor’ of the minor groove. Pullman and coworkers6 have shown that the negative electrostatic potential is greater in the AT rich region of the minor groove than GC rich regions and this provides an additional factor for AT specific minor groove binding of cationic species of molecules.
            Examples of minor groove binding drugs are netropsin7, distamycin8,9, Hoechst 3325810,11 and small indole derivatives12 (Fig. 4). DNA minor groove binding ability of small indole derivatives containing a single indole ring and a side chain of variable methyle linker length has recently been reported12,13.

Figure 4: Minor groove binders Netropsin and Distamycin. Netropsin (CPK model) is shown inside the minor groove of DNA.
            It is reported that although, the indole ring is planar, its ability to intercalate between base pairs is less. This is mainly due to less aromaticity in the ring (indole contains a six membered and a five membered ring fused together) and the absence of any positive charge in the molecule. On the other hand, the presence of methylene linker side chain enables it to orient the molecule according to the ‘shape’ of the minor groove making it a minor groove binder.

References:
1. WHO Cancer resources (2006). Retrieved from World Health Organization   http://www.who.int/cancer/publications/en/index.html
2. Kintzios S E, Barberaki M G, Plants That Fight Cancer (CRC Press) 2004.
3. Chaudhauri R P & Hargenrother P J, Curr Opin Biotech, 18 (2007) 497.
4. Canals A, Purciolas M, Aymami J & Coll M, Acta Crystallogr D Biol Crystallogr, 61 (2005) 1009.
5. Lisgarten J N, Coll M, Portugal J, Wright C W & Aymami, J Nat Struct Biol, 9 (2002) 57.
6. Lavery R & Pullman B, Int. J Quant Chem, 20 (2004) 259.
7. Patel D J & Shapiro L, J Biol Chem, 261 (1986) 1230.
8. Fagan P & Wemmer D E, J Am Chem Soc, 114 (1992) 1080.
9. Pelton J G & Wemmer D E, Proc Natl Acad Sci USA, 86 (1989) 5723.
10. Han F, Taulier N & Chalikian T V, Biochemistry 44 (2005) 9785.
11. Searle M S & Embrey K J, Nucleic Acids Res 18 (1990) 3753.
12. Pandya P, Islam M M, Kumar G S, Jayaram B & Kumar S, J Chem Sci 122 (2010) 247.
13. Gupta S P, Pandav K, Pandya P, Kumar G S, Barthwal R & Kumar S, Chem Biol Interface 1 (2011) 297.

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