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1.
Nucleic Acids Res ; 33(3): 912-21, 2005.
Artigo em Inglês | MEDLINE | ID: mdl-15703305

RESUMO

Polyamides containing an N-terminal formamido (f) group bind to the minor groove of DNA as staggered, antiparallel dimers in a sequence-specific manner. The formamido group increases the affinity and binding site size, and it promotes the molecules to stack in a staggered fashion thereby pairing itself with either a pyrrole (Py) or an imidazole (Im). There has not been a systematic study on the DNA recognition properties of the f/Py and f/Im terminal pairings. These pairings were analyzed here in the context of f-ImPyPy, f-ImPyIm, f-PyPyPy and f-PyPyIm, which contain the central pairing modes, -ImPy- and -PyPy-. The specificity of these triamides towards symmetrical recognition sites allowed for the f/Py and f/Im terminal pairings to be directly compared by SPR, CD and DeltaT (M) experiments. The f/Py pairing, when placed next to the -ImPy- or -PyPy- central pairings, prefers A/T and T/A base pairs to G/C base pairs, suggesting that f/Py has similar DNA recognition specificity to Py/Py. With -ImPy- central pairings, f/Im prefers C/G base pairs (>10 times) to the other Watson-Crick base pairs; therefore, f/Im behaves like the Py/Im pair. However, the f/Im pairing is not selective for the C/G base pair when placed next to the -PyPy- central pairings.


Assuntos
DNA/química , Imidazóis/química , Nylons/química , Pirróis/química , Pareamento de Bases , Dicroísmo Circular , Desnaturação de Ácido Nucleico , Ressonância de Plasmônio de Superfície
2.
Nucleic Acids Res ; 30(8): 1834-41, 2002 Apr 15.
Artigo em Inglês | MEDLINE | ID: mdl-11937638

RESUMO

An imidazole-containing polyamide trimer, f-ImImIm, where f is a formamido group, was recently found using NMR methods to recognize T*G mismatched base pairs. In order to characterize in detail the T*G recognition affinity and specificity of imidazole-containing polyamides, f-ImIm, f-ImImIm and f-PyImIm were synthesized. The kinetics and thermodynamics for the polyamides binding to Watson-Crick and mismatched (containing one or two T*G, A*G or G*G mismatched base pairs) hairpin oligonucleotides were determined by surface plasmon resonance and circular dichroism (CD) methods. f-ImImIm binds significantly more strongly to the T*G mismatch-containing oligonucleotides than to the sequences with other mismatched or with Watson-Crick base pairs. Compared with the Watson-Crick CCGG sequence, f-ImImIm associates more slowly with DNAs containing T*G mismatches in place of one or two C*G base pairs and, more importantly, the dissociation rate from the T*G oligonucleotides is very slow (small k(d)). These results clearly demonstrate the binding selectivity and enhanced affinity of side-by-side imidazole/imidazole pairings for T*G mismatches and show that the affinity and specificity increase arise from much lower k(d) values with the T*G mismatched duplexes. CD titration studies of f-ImImIm complexes with T*G mismatched sequences produce strong induced bands at approximately 330 nm with clear isodichroic points, in support of a single minor groove complex. CD DNA bands suggest that the complexes remain in the B conformation.


Assuntos
Pareamento Incorreto de Bases , Imidazóis/metabolismo , Nylons/química , Nylons/metabolismo , Sequência de Bases , Dicroísmo Circular , DNA/química , DNA/metabolismo , Nucleotídeos de Desoxiguanina/metabolismo , Imidazóis/química , Cinética , Conformação de Ácido Nucleico , Desnaturação de Ácido Nucleico , Ácidos Nucleicos Heteroduplexes , Ressonância de Plasmônio de Superfície , Termodinâmica , Nucleotídeos de Timina/metabolismo
3.
Nucleic Acids Res ; 32(6): 2000-7, 2004.
Artigo em Inglês | MEDLINE | ID: mdl-15064359

RESUMO

To complement available structure and binding results and to develop a detailed understanding of the basis for selective molecular recognition of T.G mismatches in DNA by imidazole containing polyamides, a full thermodynamic profile for formation of the T.G-polyamide complex has been determined. The amide-linked heterocycles f-ImImIm and f-PyImIm (where f is formamido group, Im is imidazole and Py is pyrrole) were studied by using biosensor-surface plasmon resonance (SPR) and isothermal titration calorimetry (ITC) with a T.G mismatch containing DNA hairpin duplex and a similar DNA with only Watson-Crick base pairs. Large negative binding enthalpies for all of the polyamide-DNA complexes indicate that the interactions are enthalpically driven. SPR results show slower complex formation and stronger binding of f-ImImIm to the T.G than to the match site. The thermodynamic analysis indicates that the enhanced binding to the T.G site is the result of better entropic contributions. Negative heat capacity changes for the complex are correlated with calculated solvent accessible surface area changes and indicate hydrophobic contributions to complex formation. DNase I footprinting analysis in a long DNA sequence provided supporting evidence that f-ImImIm binds selectively to T.G mismatch sites.


Assuntos
Pareamento Incorreto de Bases , Guanina/metabolismo , Imidazóis/metabolismo , Pirróis/metabolismo , Timina/metabolismo , Sequência de Bases , Sítios de Ligação , Calorimetria , DNA/química , Pegada de DNA , Imidazóis/química , Nylons/química , Pirróis/química , Ressonância de Plasmônio de Superfície , Termodinâmica
4.
J Am Chem Soc ; 127(2): 742-50, 2005 Jan 19.
Artigo em Inglês | MEDLINE | ID: mdl-15643900

RESUMO

Pyrrole (Py) and imidazole (Im) polyamides can be designed to target specific DNA sequences. The effect that the pyrrole and imidazole arrangement, plus DNA sequence, have on sequence specificity and binding affinity has been investigated using DNA melting (DeltaT(M)), circular dichroism (CD), and surface plasmon resonance (SPR) studies. SPR results obtained from a complete set of triheterocyclic polyamides show a dramatic difference in the affinity of f-ImPyIm for its cognate DNA (K(eq) = 1.9 x 10(8) M(-1)) and f-PyPyIm for its cognate DNA (K(eq) = 5.9 x 10(5) M(-1)), which could not have been anticipated prior to characterization of these compounds. Moreover, f-ImPyIm has a 10-fold greater affinity for CGCG than distamycin A has for its cognate, AATT. To understand this difference, the triamide dimers are divided into two structural groupings: central and terminal pairings. The four possible central pairings show decreasing selectivity and affinity for their respective cognate sequences: -ImPy > -PyPy- >> -PyIm- approximately -ImIm-. These results extend the language of current design motifs for polyamide sequence recognition to include the use of "words" for recognizing two adjacent base pairs, rather than "letters" for binding to single base pairs. Thus, polyamides designed to target Watson-Crick base pairs should utilize the strength of -ImPy- and -PyPy- central pairings. The f/Im and f/Py terminal groups yielded no advantage for their respective C/G or T/A base pairs. The exception is with the -ImPy- central pairing, for which f/Im has a 10-fold greater affinity for C/G than f/Py has for T/A.


Assuntos
DNA/química , Imidazóis/química , Nylons/química , Pirróis/química , Dicroísmo Circular , DNA/metabolismo , Imidazóis/metabolismo , Cinética , Conformação de Ácido Nucleico , Nylons/metabolismo , Pirróis/metabolismo , Especificidade por Substrato , Ressonância de Plasmônio de Superfície
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