The major and minor groove in duplex DNA are sites of specific molecular recognition by DNA-binding agents such as proteins, drugs and metal complexes and have functional significance. In view of this, understanding of the inherent differences in their environment and the allosteric information transfer between them induced by DNA-binding agents assumes importance. Site-specific incorporation of 5-aminodansyl-dU, (U*) in oligonucleotides d(CGCGAAU*TCGCG) and d(CGCGAATU*CGCG) leads to fluorogenic nucleic acids, in which the reporter group resides in the major groove. The fluorescent observables from such a probe are used to estimate the dielectric constant of the major groove to be approximately 55D, in comparison to the reported non polar environment of the minor groove (approximately 20D) in poly d[AT]-poly d[AT]. An exclusive minor groove event such as DNA-netropsin association can be quantitatively monitored by fluorescence of the dansyl moiety located in the major groove. This suggests existence of an information network among the two grooves. The fluorescent DNA probes as reported here may have potential applications in the study of structural polymorphisms in DNA, DNA-ligand interactions and triple helix structure.

Oligodeoxynucleotide (ODN) directed triplex formation has therapeutic importance and depends on Hoogsteen hydrogen bonds between a duplex DNA and a third DNA strand. T*A:T triplets are formed at neutral pH and C+*G:C are favoured at acidic pH. It is demonstrated that spermine conjugation at N4 of 5-Me-dC in ODNs 1-5 (sp-ODNs) imparts zwitterionic character, thus reducing the net negative charge of ODNs 1-5. sp-ODNs form triplexes with complementary 24mer duplex 8:9 show foremost stability at neutral pH 7.3 and decrease in stability towards lower pH, unlike the normal ODNs where optimal stability is found at an acidic pH 5.5. At pH 7.3, control ODNs 6 and 7 carrying dC or 5-Me-dC, respectively, do not show any triple helix formation. The stability order of triplex containing 5-Me-dC-N4-(spermine) with normal and mismatched duplex was found to be X*G:C approximately X*A:T > X*C:G > X*T:A. The hysteresis curve of sp-ODN triplex 3*8:9 indicated a better association with complementary duplex 8:9 as compared to unmodified ODN 6 in triplex 6*8:9. pH-dependent UV difference spectra suggest that N3 protonation is not a requirement for triplex formation by sp-ODN and interstrand interaction of conjugated spermine more than compensates for loss in stability due to absence of a single Hoogsteen hydrogen bond. These results may have importance in designing oligonucleotides for antigene applications.

The synthesis of mixed backbone oligodeoxynucleotides (18-mers) consisting of positively charged guanidinium linkages along with negatively charged phosphodiester linkages is carried out. The use of a base labileprotecting group for guanidinium linkage offers a synthetic strategy similar to standard oligonucleotide synthesis. The nuclease resistance of the oligodeoxyribonucleotides capped with guanidinium linkages at 5 and 3 ends are reported. The hybridization properties and sequence specificity of binding of these deoxynucleic guanidine͞DNA chimeras with complementary DNA or RNA are described.The use of antisense oligodeoxyribonucleotides (ODNs) to regulate gene products requires the development of modified ODNs possessing the properties of enhanced cellular uptake, nuclease resistance, and sequence specific hybridization to complementary RNAs. Numerous DNA structural analogues with modified heterocycle, sugar, and phosphodiester backbone moieties have been synthesized (1-3). Substantial progress has been made toward successful backbone modifications by using phosphorus and non-phosphorus groups (4, 5). A number of modifications or replacements of phosphodiester linkages such as 2Ј-fluoro-N-3Ј-P5Ј-phosphoramidates (6), 3Ј-thioformacetals (7, 8), 2Ј-O-Me methylene(methylimino) (9), 2Ј-O-Me amide (10), 2Ј-O-methylribonucleoside methylphosphonate (11), and peptide nucleic acid (PNA) (12) have been shown to complement with DNA and RNA with similar or higher stability while maintaining the sequence specificity. Except for the 2Ј-fluoro-N3Ј-P5Ј-phosphoramidates, these analogues are neutral and thus eliminate the electrostatic repulsion of negative charges present in natural DNA and RNA. An alternative approach, involves replacement of anionic phosphodiester groups by cationic linkages (13-17) or the use of oligonucleotides conjugated with positively charged groups to provide zwitterionic DNA analogues (18)(19)(20). These ODNs show increased binding with complementary DNA or RNA. Conceptually, replacement of anionic phosphodiester linkage by neutral or positively charged linkages can modulate the net charge of antisense ODN complex and thereby may enhance its antisense properties (4).Our ongoing research in this area is focused on the development of deoxynucleic guanidine (DNG) in which the negatively charged OOO(PO 2 Ϫ )OOO backbone of DNA is replaced by positively charged, achiral ONHOC(ANH 2 ϩ )ONH O linkage (15-17) to provide very stable complexes (21, 22). As DNG is positively charged, it binds effectively to target DNA or RNA because the repulsive electrostatic effects in double-stranded DNA (dsDNA) would be replaced by attractive electrostatic interactions in DNG:DNA or DNG:RNA duplexes. On the other hand, if electrostatic binding between polycationic and polyanionic structures becomes more significant than the specific interactions between heterocyclic bases, then binding becomes nonspecific and independent of complementary base pairing. To overcome this possible limitation, we propose to synthesize m...

DNA duplex recognition by macromolecules (proteins, enzymes) and small molecules (drugs, metal complexes) occurs in the major or minor grooves of DNA via hydrogen bonding and electrostatic or hydrophobic interactions, whose strengths depend on the medium. It is therefore important to understand the local environments of the major/minor grooves of DNA. It is known that small molecules bind in the minor groove that have nonpolar character (ε = 22) and are organic-like. By employing fluorescent oligonucleotides, we demonstrated recently that the major groove of DNA is more polar than the minor groove (ε = 55) and the DNA double helix has different polarity in its two grooves. Our experimentally measured values were recently validated independently by two theoretical groups, employing calculations by two different methodologies. In this paper, a hitherto unknown property of DNA double helixsequence dependent local polarity or microenvironmentis experimentally demonstrated. This is shown by fluorescence experiments using oligonucleotides containing the modified base 5-amino-dU in place of dT and the polarity varied between ε ≈ 40 and 60, in different sequences. This is the first experimental demonstration of sequence-dependent microenvironmental effects in DNA in solution, although structural effects are well-known by X-ray data. The technique used has a good potential to investigate differing microenvoronmental effects in various secondary structures, DNA polymorphs, and chemically modified DNA and their hybrids.

Our initial structure-activity relationship studies on 7-methoxy-4-morpholino-benzothiazole derivatives featured by aryloxy-2-methylpropanamide moieties at the 2-position led to identification of compound 25 as a potent and selective A adenosine receptor (AAdoR) antagonist with reasonable ADME and pharmacokinetic properties. However, poor intrinsic solubility and low to moderate oral bioavailability made this series unsuitable for further development. Further optimization using structure-based drug design approach resulted in discovery of potent and selective adenosine A receptor antagonists bearing substituted 1-methylcyclohexyl-carboxamide groups at position 2 of the benzothiazole scaffold and endowed with better solubility and oral bioavailability. Compounds 41 and 49 demonstrated a number of positive attributes with respect to in vitro ADME properties. Both compounds displayed good pharmacokinetic properties with 63% and 61% oral bioavailability, respectively, in rat. Further, compound 49 displayed oral efficacy in 6-OHDA lesioned rat model of Parkinson diseases.

No abstract

No abstract

A fully automated solid-phase synthetic procedure for incorporation of positively charged guanidinium linkages into otherwise neutral PNA sequences has been employed. These DNG/PNA chimeras form [(DNG/ PNA) 2 ‚DNA] triplexes upon binding to single strand or duplex DNA (with accompanying D-loop for the latter). The [(DNG/PNA) 2 ‚DNA] triplexes of DNG/PNA T 10 , with DNA dA 10 , are more stable than DNA‚ DNA triplexes [(T 10 ) 2 ‚dA 10 ]. The binding of DNG/PNA chimera with guanidinium linkages on both ends, with single strand length-matched complementary DNA under thermal melt conditions and with longer double strand DNA under isothermal conditions is sequence specific. The association process of DNG/PNA chimera with single strand DNA and strand invasion of longer ds-DNA is faster than the association of PNA, with the same DNA targets, as evident by thermal hysteresis and gel retardation under isothermal conditions. The sequence specific and faster strand invasion of DNG/PNA may extend the potential utility of PNA in diagnostics, biomolecular probes, and antisense/antigene therapeutics.