Dataset Information

CSI-FID: high throughput label-free detection of DNA binding molecules.

ABSTRACT: Determining the sequence specifity of DNA binding molecules is a non-trivial task. Here we describe the development of a platform for assaying the sequence specificity of DNA ligands using label free detection on high density DNA microarrays. This is achieved by combining Cognate Site Identification (CSI) with Fluorescence Intercalation Displacement (FID) to create CSI-FID. We use the well-studied small molecule DNA ligand netropsin to develop this high throughput platform. Analysis of the DNA binding properties of protein- and small molecule-based libraries with CSI-FID will advance the development of genome-anchored molecules for therapeutic purposes.

SUBMITTER: Hauschild KE

PROVIDER: S-EPMC2760306 | biostudies-literature | 2009 Jul

REPOSITORIES: biostudies-literature

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CSI-FID: high throughput label-free detection of DNA binding molecules.

Hauschild Karl E KE Stover James S JS Boger Dale L DL Ansari Aseem Z AZ

Bioorganic & medicinal chemistry letters 20090503 14

Determining the sequence specifity of DNA binding molecules is a non-trivial task. Here we describe the development of a platform for assaying the sequence specificity of DNA ligands using label free detection on high density DNA microarrays. This is achieved by combining Cognate Site Identification (CSI) with Fluorescence Intercalation Displacement (FID) to create CSI-FID. We use the well-studied small molecule DNA ligand netropsin to develop this high throughput platform. Analysis of the DNA b ...[more]

PMID: 19435662

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Project description:The identification and characterization of binding sites of DNA-binding molecules, including transcription factors (TFs), is a critical problem at the interface of chemistry, biology and molecular medicine. The Cognate Site Identification (CSI) array is a high-throughput microarray platform for measuring comprehensive recognition profiles of DNA-binding molecules. This technique produces datasets that are useful not only for identifying binding sites of previously uncharacterized TFs but also for elucidating dependencies, both local and nonlocal, between the nucleotides at different positions of the recognition sites. We have developed a regression tree technique, CSI-Tree, for exploring the spectrum of binding sites of DNA-binding molecules. Our approach constructs regression trees utilizing the CSI data of unaligned sequences. The resulting model partitions the binding spectrum into homogeneous regions of position specific nucleotide effects. Each homogeneous partition is then summarized by a position weight matrix (PWM). Hence, the final outcome is a binding intensity rank-ordered collection of PWMs each of which spans a different region in the binding spectrum. Nodes of the regression tree depict the critical position/nucleotide combinations. We analyze the CSI data of the eukaryotic TF Nkx-2.5 and two engineered small molecule DNA ligands and obtain unique insights into their binding properties. The CSI tree for Nkx-2.5 reveals an interaction between two positions of the binding profile and elucidates how different nucleotide combinations at these two positions lead to different binding affinities. The CSI trees for the engineered DNA ligands exhibit a common preference for the dinucleotide AA in the first two positions, which is consistent with preference for a narrow and relatively flat minor groove. We carry out a reanalysis of these data with a mixture of PWMs approach. This approach is an advancement over the simple PWM model and accommodates position dependencies based on only sequence data. Our analysis indicates that the dependencies revealed by the CSI-Tree are challenging to discover without the actual binding intensities. Moreover, such a mixture model is highly sensitive to the number and length of the sequences analyzed. In contrast, CSI-Tree provides interpretable and concise summaries of the complete recognition profiles of DNA-binding molecules by utilizing binding affinities.

Dataset Information

CSI-FID: high throughput label-free detection of DNA binding molecules.

Publications

CSI-FID: high throughput label-free detection of DNA binding molecules.

Similar Datasets

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