Modifying the N-terminus of polyamides: PyImPyIm has improved sequence specificity over f-ImPyIm

Brown, Toni; Mackay, Hilary; Turlington, Mark; Sutterfield, Arden; Smith, Traci; Sielaff, Alan; Westrate, Laura M.; Bruce, Carolyn; Kluza, Jérôme; O’Hare, Caroline; Nguyen, Binh; Wilson, W. David; Hartley, John A.; Lee, Moses

doi:10.1016/j.bmc.2008.03.008

Cited by 15 publications

(41 citation statements)

References 44 publications

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“…The nitro group of imidazolecarboxamide ( 4 ) was reduced using standard Pd-C catalyzed hydrogenation conditions as reported previously [10] and the resulting amine was reacted with the relevant commercially available nitrobenzoyl chloride using standard Schotten-Bauman coupling conditions [11]. The final imidazole heterocyle was added in the same manner as described in previously reports [9], and the formylation step was performed using the published procedure [10]. All compounds were obtained in good yield and pure according to TLC and 1 H-NMR analyses.…”

Section: Resultsmentioning

confidence: 99%

“…With regard to f-ImPyIm ( 3 ), similar results are observed for both compound 2c and f-Im-Py-Im ( 3 ), with respect to binding stoichiometry and CD response. Both compounds produce a CD ellipticity of ~35 mdeg for the cognate sequence (A CGCG T) and a response of ~ 13 mdeg for the non-cognate (A AATT T) DNA sequence [10]. …”

Section: Resultsmentioning

confidence: 99%

“…Thermal denaturation data was obtained using a Cary 100 BioMelt (Varian) spectrophotometer with DNA (1 μM) in PO 4 0 (10 mM phosphate buffer, containing 10 mM Na + , and 1 mM EDTA, pH 6.2), and compounds 2a–c (3 μM) , using the procedure previously reported [8,10b]. Thermal melts were obtained using both DNA’s (A CGCG T and A AATT T) and each of the three polyamide compounds ( 2a–c ).…”

Section: Methodsmentioning

confidence: 99%

“…Circular Dichroism (CD) titration studies were performed on an OLIS DSM20 spectropolarimeter, using a 1 mm path-length cuvette using the previously published procedure [10]. Experiments were carried out using PO 4 5 (10 mM phosphate buffer, containing 50 mM Na + , and 1 mM EDTA, pH 6.2), and 160 μL of a 9 μM DNA solution ( ACGCGT and AAATTT ) was titrated with 1 mol equivalents of the poly-amide f-Im-Phenyl-Im 2a–c , past the point of saturation.…”

Section: Methodsmentioning

confidence: 99%

“…ITC analysis was performed using a VP-ITC micromalorimeter (MicroCal) using the previously reported procedure [8,10b]. Compounds 2a–c were dissolved in PO 4 5 and the instrument equilibrated at 25 ºC.…”

Section: Methodsmentioning

confidence: 99%

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Polyamide Curvature and DNA Sequence Selective Recognition: Use of 4-Aminobenzamide to Adjust Curvature

Lajiness¹,

Sielaff²,

Mackay³

et al. 2009

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Imidazole and pyrrole-containing polyamides belong to an important class of compounds that can be designed to target specific DNA sequences, and they are potentially useful in applications of controlling gene expression. The extent of polyamide curvature is an important consideration when studying the ability of such compounds to bind in the minor groove of DNA. The current study investigates the importance of curvature using polyamides of the form f-Im-Phenyl-Im, in which the imidazole heterocycles are placed in ortho-, meta-, and para-configurations of the phenyl moiety. The synthesis and biophysical evaluation of each compound binding to its cognate DNA sequence (5′-ACGCGT-3′) and a negative control sequence (5′-AAATTT-3′) is reported, along with their comparison to the parent binder, f-Im-Py-Im (3). ACGCGT is a medicinally significant sequence present in the MluI cell-cycle box (MCB) transcriptional element found in the promoter of a gene associated with cell division. The results demonstrated that the para-derivative has the greatest affinity for its cognate sequence, as indicated via thermal denaturation, CD, ITC, SPR analyses, and DNase I footprinting. ITC studies showed that binding of the para-isomer (2c) to ACGCGT was significantly more exothermic than binding to AAATTT. In contrast, no heat change was observed for binding of the meta- (2b) and ortho- (2a) isomers to both DNAs, due to low binding affinities. This is consistent with results from SPR studies, which indicate that the para-derivative binds in a 2:1 fashion to ACGCGT and binds weakly to ACCGGT (K = 1.8 × 106 and 4.0 × 104 M−1, respectively). Interestingly, it binds in a 1:1 fashion to AAATTT (K = 5.4 × 105 M−1). The meta-compound does not bind to any sequence. The para-derivative also was the only compound to show an induced peak via CD at 330 nm, indicative of minor groove binding, and produced a ΔTm value of 5.8 ºC. Molecular modeling experiments have been performed to determine the shape differences between the three compounds, and the results indicate that the para-derivative 2c has a closest curvature to previously synthesized polyamides. DNase I footprinting studies confirmed earlier observations that only the para-derivative 2c produced a footprint with ACGCGT (1 μM) and no significant footprint was observed at any sites examined for meta-2b and ortho-2a analogs up to 40 μM. The results of these studies suggest that the shape of the ortho- and meta- derivatives is too curved to match the curvature of the DNA minor groove to facilitate binding. The para-derivative gives the highest binding affinity in the series and the results illustrate that 4-aminobenzamide is a reasonable substitute for 4-aminopyrrole-2-carboxylate.

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Section: Resultsmentioning

confidence: 99%

Section: Resultsmentioning

confidence: 99%

Section: Methodsmentioning

confidence: 99%

Section: Methodsmentioning

confidence: 99%

Section: Methodsmentioning

confidence: 99%

See 3 more Smart Citations

Polyamide Curvature and DNA Sequence Selective Recognition: Use of 4-Aminobenzamide to Adjust Curvature

Lajiness¹,

Sielaff²,

Mackay³

et al. 2009

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DNA‐Binding Properties of New Fluorescent AzaHx Amides: Methoxypyridylazabenzimidazolepyrroleimidazole/pyrrole

et al. 2018

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DNA minor groove binding polyamides have been extensively developed to control abnormal gene expression. The establishment of novel, inherently fluorescent 2-(p-anisyl)benzimidazole (Hx) amides has provided an alternative path for studying DNA binding in cells by direct observation of cell localization. Because of the 2:1 antiparallel stacking homodimer binding mode of these molecules to DNA, modification of Hx amides to 2-(p-anisyl)-4-azabenzimidazole (AzaHx) amides has successfully extended the DNA-recognition repertoire from central CG [recognized by Hx-I (I=N-methylimidazole)] to central GC [recognized by AzaHx-P (P=N-methylpyrrole)] recognition. For potential targeting of two consecutive GG bases, modification of the AzaHx moiety to 2- and 3-pyridyl-aza-benzimidazole (Pyr-AzaHx) moieties was explored. The newly designed molecules are also small-sized, fluorescent amides with the Pyr-AzaHx moiety connected to two conventional five-membered heterocycles. Complementary biophysical methods were performed to investigate the DNA-binding properties of these molecules. The results showed that neither 3-Pyr-AzaHx nor 2-Pyr-AzaHx was able to mimic I-I=N-methylimidazole-N-methylimidazole to target GG dinucleotides specifically. Rather, 3-Pyr-AzaHx was found to function like AzaHx, f-I (f=formamide), or P-I as an antiparallel stacked dimer. 3-Pyr-AzaHx-PI (2) binds 5'-ACGCGT'-3' with improved binding affinity and high sequence specificity in comparison to its parent molecule AzaHx-PI (1). However, 2-Pyr-AzaHx is detrimental to DNA binding because of an unfavorable steric clash upon stacking in the minor groove.

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Grading the commercial optical biosensor literature—Class of 2008: ‘The Mighty Binders’

Rich

Myszka

2009

J of Molecular Recognition

139

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Optical biosensor technology continues to be the method of choice for label-free, real-time interaction analysis. But when it comes to improving the quality of the biosensor literature, education should be fundamental. Of the 1413 articles published in 2008, less than 30% would pass the requirements for high-school chemistry. To teach by example, we spotlight 10 papers that illustrate how to implement the technology properly. Then we grade every paper published in 2008 on a scale from A to F and outline what features make a biosensor article fabulous, middling or abysmal. To help improve the quality of published data, we focus on a few experimental, analysis and presentation mistakes that are alarmingly common. With the literature as a guide, we want to ensure that no user is left behind.

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Modifying the N-terminus of polyamides: PyImPyIm has improved sequence specificity over f-ImPyIm

Cited by 15 publications

References 44 publications

Polyamide Curvature and DNA Sequence Selective Recognition: Use of 4-Aminobenzamide to Adjust Curvature

Polyamide Curvature and DNA Sequence Selective Recognition: Use of 4-Aminobenzamide to Adjust Curvature

DNA‐Binding Properties of New Fluorescent AzaHx Amides: Methoxypyridylazabenzimidazolepyrroleimidazole/pyrrole

Grading the commercial optical biosensor literature—Class of 2008: ‘The Mighty Binders’

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