Determination of Binding Constant of DNA-binding Drug to Target DNA by Surface Plasmon Resonance Biosensor Technology

Abstract

The experimental determination of the binding constant of a drug for its target molecule is of considerable importance. It is a basic experimental parameter in a variety of studies, such as the prediction of drug efficiency, or in the pharmacokinetic drug interaction. DNA-binding drugs have been reported to be able to interfere in a sequence dependent manner with biological functions such as topoisomerase activity, restriction of enzyme cleavage of DNA, protein-DNA interactions and the activity of transcription factors, leading to alteration of gene expression. This effect could have important practical application in the experimental therapy of human pathologies, including neoplastic diseases and viral, or microbial infections. The assessment of the biological activity of DNA-binding drugs by polymerase chain reaction, footprinting, gel retardation and in vitro transcription studies was recently reported. However, most of these techniques are steady-state methodologies and therefore are not suitable for an easy determination of the binding activity of DNA-binding drugs to target DNA and the stability of drugs-DNA complexes. Direct real-time observation and measurement of the interaction between DNA-binding drug and target DNA sequence is a subject of interest for drug discovery and development. The recent development of biosensors, based on surface plasmon resonance (SPR) technology, enables monitoring of a variety of biospecific interactions of DNA-binding drugs with target DNA elements in real-time. The present review is designed to indicate the theoretical background of SPR-based biosensor technology as well as to present the great variety of measurements and modes of interaction kinetics that can be performed with these techniques. In addition, some of the most recent studies in determining the binding constant and stoichiometry of DNA-binding drugs to target DNA with SPR technology are reviewed and the available theoretical aspects necessary for the comprehension of the experiments are provided.