Journal article
Rigid Double-Stranded DNA Linkers for Single Molecule Enzyme-Drug Interaction Measurements Using Molecular Recognition Force Spectroscopy
Langmuir, Vol.36(15), pp.4174-4183
04/21/2020
DOI: 10.1021/acs.langmuir.9b03495
PMID: 32233509
Abstract
Single-molecule studies can reveal the distribution of states and interactions between ligand-enzyme complexes not accessible for most studies that measure a large ensemble average response of many molecules. Furthermore, in some biological applications, the information regarding the outliers, not the average of measured properties, can be more important. The high spatial and force resolution provided by atomic force microscopy (AFM) under physiological conditions has been utilized in this study to quantify the force-distance relations of enzyme-drug interactions. Different immobilization techniques of the protein to a surface and the drug to AFM tip were quantitatively compared to improve the accuracy and precision of the measurement. Protein that is directly bound to the surface, forming a monolayer, was compared to enzyme molecules bound to the surface with rigid double-stranded (ds) DNA spacers. These surfaces immobilization techniques were studied with the drug bound directly to the AFM tip and drug bound via flexible poly(ethylene glycol) and rigid dsDNA linkers. The activity of the enzyme was found to be not significantly altered by immobilization methods relative to its activity in solution. The findings indicate that the approach for studying drug-enzyme interaction based on rigid dsDNA linker on the surface and either flexible or rigid linker on the tip affords straightforward, highly specific, reproducible, and accurate force measurements with a potential for single-molecule level studies. The method could facilitate in-depth examination of a broad spectrum of biological targets and potential drugs.
Details
- Title: Subtitle
- Rigid Double-Stranded DNA Linkers for Single Molecule Enzyme-Drug Interaction Measurements Using Molecular Recognition Force Spectroscopy
- Creators
- Thiranjeewa I Lansakara - Department of Chemistry, University of Iowa, Iowa City, Iowa 52242, United StatesHolly S Morris - Department of Chemistry, University of Iowa, Iowa City, Iowa 52242, United StatesPriyanka Singh - Department of Chemistry, University of Iowa, Iowa City, Iowa 52242, United StatesAmnon Kohen - Department of Chemistry, University of Iowa, Iowa City, Iowa 52242, United StatesAlexei V Tivanski - Department of Chemistry, University of Iowa, Iowa City, Iowa 52242, United States
- Resource Type
- Journal article
- Publication Details
- Langmuir, Vol.36(15), pp.4174-4183
- DOI
- 10.1021/acs.langmuir.9b03495
- PMID
- 32233509
- ISSN
- 0743-7463
- eISSN
- 1520-5827
- Grant note
- DOI: 10.13039/100000165, name: Division of Chemistry, award: CHE-1149023; DOI: 10.13039/100000002, name: National Institutes of Health, award: R01GM65368
- Language
- English
- Date published
- 04/21/2020
- Academic Unit
- Chemistry
- Record Identifier
- 9984216676202771
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