Quantitative characterization of conformational-specific protein-DNA binding using a dual-spectral interferometric imaging biosensor

Nanoscale
Xirui ZhangM Selim Unlü

Abstract

DNA-binding proteins play crucial roles in the maintenance and functions of the genome and yet, their specific binding mechanisms are not fully understood. Recently, it was discovered that DNA-binding proteins recognize specific binding sites to carry out their functions through an indirect readout mechanism by recognizing and capturing DNA conformational flexibility and deformation. High-throughput DNA microarray-based methods that provide large-scale protein-DNA binding information have shown effective and comprehensive analysis of protein-DNA binding affinities, but do not provide information of DNA conformational changes in specific protein-DNA complexes. Building on the high-throughput capability of DNA microarrays, we demonstrate a quantitative approach that simultaneously measures the amount of protein binding to DNA and nanometer-scale DNA conformational change induced by protein binding in a microarray format. Both measurements rely on spectral interferometry on a layered substrate using a single optical instrument in two distinct modalities. In the first modality, we quantitate the amount of binding of protein to surface-immobilized DNA in each DNA spot using a label-free spectral reflectivity technique that accuratel...Continue Reading

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Citations

May 1, 2021·International Journal of Molecular Sciences·Thomas CarzanigaTommaso Bellini
Aug 30, 2017·ACS Applied Materials & Interfaces·Nhu Hoa Thi TranHeongkyu Ju
Aug 2, 2020·Biophysical Journal·Luka VanjurMarco Buscaglia

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