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Differential Scanning Calorimetry to Study DNA Aptamer-Thermolabile Ligand Interaction

Published: June 29, 2023

Abstract

Source: Mittermaier, A. K., et al. Measuring Biomolecular DSC Profiles with Thermolabile Ligands to Rapidly Characterize Folding and Binding Interactions. J. Vis. Exp. (2017)

This video demonstrates differential scanning calorimetry — a technique to study the interaction between a biomolecule and a ligand. The binding of a DNA aptamer and a thermolabile ligand is studied by applying repeated heating and cooling cycles to the sample and obtaining scans for thermodynamic analysis.

Protocol

1. Sample Preparation Purify the desired biomolecule. NOTE: This protocol uses purchased cocaine-binding DNA aptamer MN4 after exchanging against 2 M NaCl three times followed by three rounds of deionized water using a centrifugal filter with a 3 kDa molecular weight cut-off membrane. Synthesize and purify, or purchase the desired thermolabile ligand. NOTE: MN4 binds the thermolabile ligand cocaine. MN4 also binds quinine, which is use…

Disclosures

The authors have nothing to disclose.

Materials

Sodium chloride Chem Impex #00829
Sodium phosphate monobasic dihydrate Sigma Aldrich 71502
Sodium phosphate dibasic Sigma Aldrich S9763
Deioinized water for molecular biology Millipore H20MB1001
0.2-micron sterile syringe filters VWR CA28145-477
3 kDa centrifugal filters Millipore UFC900324
Dialysis tubing 0.5-1.0 kDa cutoff Spectrum Laboratories 131048
Silicon tubing VWR 89068-474
Plastic DSC flange caps TA Instruments 6111
DNA aptamer MN4 Integrated DNA Technologies https://www.idtdna.com/site/order/menu
Cocaine Sigma Aldrich C008
Quinine Sigma Aldrich 22620
NanoDSC-III microcalorimeter TA Instruments http://www.tainstruments.com/nanodsc/
DSCRun software TA Instruments http://www.tainstruments.com/support/software-downloads-support/instruments-by-software/
NanoAnalyze software TA Instruments http://www.tainstruments.com/support/software-downloads-support/instruments-by-software/
Contrad-70 VWR 89233-152

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Cite This Article
Differential Scanning Calorimetry to Study DNA Aptamer-Thermolabile Ligand Interaction. J. Vis. Exp. (Pending Publication), e21450, doi: (2023).

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