Published October 14, 2020 | Version v1
Dataset Open

LILBID laser dissociation curves: a mass spectrometry-based method for the quantitative assessment of dsDNA binding affinities

  • 1. Goethe University Frankfurt

Description

The data and data analysis scripts in this dataset are referenced in the manuscript, "LILBID laser dissociation curves: a mass spectrometry-based method for the quantitative assessment of dsDNA binding affinities", which is in preparation for publication. The contents of this dataset are as follows:

1) raw data from UV melting curves
2) settings, concentrations, and both raw and processed data from ITC experiments
3) raw spectrum and imaging data from qLILBID experiments
4) programming scripts used to process the qLILBID data

Notes on the ITC data:
The iTC200 microcalorimeter (Malvern Panalytical, Malvern, UK) used in the ITC experiments produces .itc files to be opened and analyzed in Origin (Originlab, Northampton, MA, US) using an add-on. The resulting Origin files, including data interpretation and figures, are provided here. Raw data and interpreted data have been gathered from the .itc files and the Origin files and assembled into tab-separated .dat files, so that the data are also accessible to users who do not have Origin.

The Origin files can be understood as follows. After data collection, the ITC raw data are loaded into the Origin-based software. Initially, the baseline is created (Data1Coeff worksheet) and the data plotted in µcal/second as a function of time (minutes), shown in the mRawITC (graph) and the Data1RAW (data) windows. The peaks are integrated (area in µcal) and then plotted in units of kcal/mole of injectant as a function of molar ratio (injected ligand per molecule in the cell), shown in the DeltaH window. The first injection is negligible and therefore always deleted. According to the data points in the DeltaH window, a curve is fitted to obtain the molar ratio (N), Ka, ΔH and ΔS, the data is shown in the Data1 worksheet. The Data1 worksheet hereby contains the following information: DH: heat change resulting from the given injection (µcal/injection); INJV: volume of the injection; Xt: concentration of injected ligand in the cell before next injection; Mt: concentration of molecule in the cell before next injection; volume corrected; XMt: molar ratio of ligand per molecule in the cell after the injection as displayed in the DeltaH window; NDH: Normalized DH in kcal/moles of injectant as displayed in the DeltaH window, Fit: data points of the fitted curve. In the end, the results are presented in the ITCFINAL window (final figure). Additional information can be found in the MicroCal iTC200 System User Manual.

The same data labeling system has been used for the tab-separated .dat files.

Notes

Acknowledgments: We thank Heidi Zetsche and Yvonne Becker for their input with the UV measurements. P.Y. was supported by the DFG (German Research Foundation), Collaborative Research Center 807, "Transport and Communication across Biological Membranes." N.M. received funding by the Cluster of Excellence Frankfurt (Macromolecular Complexes) the European Research Council under the European Union's Seventh Framework Programme (FP7/2007−2013)/ERC Grant agreement n° 337567 and received funding from the DFG via a Heisenberg professorship. C.I. is supported by the DFG through grant Wo901/7-1 to J.W. The ITC instrument was acquired with funds from the DFG-supported CRC 902 "Molecular Principles of RNA-based Regulation."

Files

UV melting 10_Amer 19.07.2018_ 1.txt

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