ABSTRACT
The effect of amino groups attached at positions 2 and 8 of the hypoxanthine moiety in the structure, reactivity and stability of DNA duplexes and triplexes is studied by means of quantum mechanical calculations, as well as extended molecular dynamics (MD) and thermodynamic integration (MD/TI) simulations. Theoretical estimates of the change in stability related to 2'-deoxyguanosine (G) --> 2'-deoxyinosine (I) --> 8-amino-2'-deoxyinosine (8AI) mutations have been experimentally verified, after synthesis of the corresponding compounds. An amino group placed at position 2 stabilizes the duplex, as expected, and surprisingly also the triplex. The presence of an amino group at position 8 of the hypoxanthine moiety stabilizes the triplex but, surprisingly, destabilizes the duplex. The subtle electronic redistribution occurring upon the introduction of an amino group on the purine seems to be responsible for this surprising behavior. Interesting 'universal base' properties are found for 8AI.
Subject(s)
DNA/chemistry , DNA/metabolism , Inosine/metabolism , Base Sequence , Computer Simulation , DNA/chemical synthesis , DNA/genetics , Hydrogen Bonding , Inosine/chemistry , Inosine/genetics , Models, Molecular , Molecular Structure , Mutation , Nucleic Acid Conformation , Nucleic Acid Denaturation , Oligodeoxyribonucleotides/chemical synthesis , Oligodeoxyribonucleotides/chemistry , Oligodeoxyribonucleotides/genetics , Oligodeoxyribonucleotides/metabolism , Quantum Theory , ThermodynamicsABSTRACT
A DNA-triplex stabilizing purine (8-aminoguanine) is designed based on molecular modeling and synthesized. The substitution of guanine by 8-aminoguanine largely stabilizes the triplex both at neutral and acidic pH, as suggested by molecular dynamics and thermodynamic integration calculations, and demonstrated by melting experiments. NMR experiments confirm the triplex-stabilizing properties of 8-aminoguanine and demonstrate that few changes are found in the structure of the triplex due to the presence of this modified base.
Subject(s)
DNA/chemistry , Guanine/analogs & derivatives , Guanine/chemistry , Nucleic Acid Conformation , Cytosine/chemistry , DNA/genetics , Magnetic Resonance Spectroscopy/methods , Models, Molecular , Mutation , Nucleic Acid Denaturation , Structure-Activity Relationship , ThermodynamicsABSTRACT
The synthesis of parallel hairpins carrying 8-aminopurines is described. These hairpins have a high affinity for specific polypyrimidine sequences resulting in the formation of very stable triplexes.
Subject(s)
Oligodeoxyribonucleotides/chemistry , Purines/chemistry , Base Sequence , Models, Molecular , Nucleic Acid Conformation , Nucleic Acid Denaturation , Oligodeoxyribonucleotides/chemical synthesis , ThermodynamicsABSTRACT
Analysis of the Hoogsteen base pairs of guanine and 8-aminoguanine with cytosine in the gase phase shows a strong stabilization of the 8-aminoguanine.cytosine base pair due to the formation of an extra H-bond. Melting profiles of oligonucleotides carrying 8-amino-2'-deoxyguanosine confirm the triple helix stabilization properties of 8-aminoguanine.
Subject(s)
Deoxyguanosine/chemistry , Guanine/analogs & derivatives , Oligonucleotides/chemistry , Drug Stability , Guanine/chemistry , Hydrogen BondingABSTRACT
The incorporation of 5-azacytosine residues into DNA causes potent inhibition of DNA (Cytosine-C5) methyltransferases. The synthesis of oligodeoxyribonucleotides incorporating single or multiple 5-aza-2'-deoxycytidine residues at precise sites was undertaken to generate an array of sequences containing the reactive 5-azacytosine base as specific target sites for enzymatic methylation. Preparation of these modified oligonucleotides requires the use of 2-(p-nitrophenyl)ethyloxycarbonyl (NPEOC) groups for the protection of exocyclic amino functions. These groups are removed under mild conditions, thus avoiding conventional protocols that are detrimental to the integrity of the 5-azacytosine ring.
Subject(s)
Cytosine/analogs & derivatives , DNA (Cytosine-5-)-Methyltransferases/antagonists & inhibitors , Oligodeoxyribonucleotides, Antisense/chemical synthesis , Oligodeoxyribonucleotides, Antisense/pharmacology , Base Sequence , Indicators and Reagents , Oligodeoxyribonucleotides, Antisense/chemistryABSTRACT
Oligonucleotides containing the photoreactive nucleosides 2-azido-2'-deoxyinosine and 8-azido-2'-deoxyadenosine have been prepared using protected 2-fluoro-2'-deoxyinosine and 8-bromo-2'-deoxyadenosine phosphoramidites. After the assembly of the oligonucleotides, the nucleoside derivatives are converted to the corresponding azido derivatives by treatment with lithium azide in dry DMF. Deprotection of oligonucleotides carrying these azidonucleosides is performed with concentrated ammonia at room temperature.
Subject(s)
Adenosine/analogs & derivatives , Azides/chemistry , Inosine/analogs & derivatives , Oligodeoxyribonucleotides/chemical synthesis , Adenosine/chemistry , Ammonia , Deoxyadenosines/chemistry , Inosine/chemistry , NucleosidesABSTRACT
A novel approach for the preparation of oligonucleotides carrying amino groups at the 3'-end is described. Several CPG supports having aminoalkyl groups and 3'-amino-2',3'-dideoxynucleosides linked through base-labile carbamate linkages such as 2-(2-nitrophenyl)ethoxycarbonyl and fluorenylmethoxycarbonyl were prepared using two different strategies. These supports are compatible to the standard solid phase phosphite-triester methodology and yield oligonucleotides containing amino groups at the 3'-end. Several properties of the 3'-amino oligonucleotides, such as nuclease resistance, hybridization, and preparation of oligonucleotide conjugates are discussed.