DOI: 10.5061/dryad.t76hdr8df
title: Oligomeric state and ATP binding to <em>E. coli</em> Rho measured by native MS

description: Mass photometry and mass spectrometry data of E. coli Rho helicase over a
 range of concentrations, and in the presence of varying ATP
 concentrations. Python script for fitting population data. Data show that
 Rho adopts multiple oligomeric states over the range of sampled conditions
 and that ATP binds variably to these oligomeric states.
description: # Oligomeric state and ATP binding to *E. coli* Rho measured by native MS
 Dataset DOI:
 [10.5061/dryad.t76hdr8df](https://doi.org/10.5061/dryad.t76hdr8df) ##
 Description of the data and file structure We have submitted all native
 mass spectra appearing in the paper, ratiometric contrast counts from mass
 photometry experiments, and A280 traces from analytical size-exclusion
 chromatography experiments. Scripts used to analyze data are included in
 their respective directories. The zip file `Billings_20251010.zip`
 contains the following. ### Figure1B #### 20241209_10uMRho_IST100.txt *
 *Header*: 7 lines summarizing acquisition parameters and the file
 structure * *Mass*: Mass-to-charge ratio of ions detected (m/z, Th) *
 *Intensity*: Measure of ion abundance (intensity units) ### Figure1C ####
 2xTBS_SEC.csv * *mL-100nM*: Running volume of column injected with 100
 nanomolar Rho (mL) * *nI-100nM*: Normalized intensity of A280 trace
 (unitless) * *mL-1uM*: Running volume of column injected with 1 micromolar
 Rho (mL) * *nI-1uM*: Normalized intensity of A280 trace (unitless) *
 *mL-10uM*: Running volume of column injected with 10 micromolar Rho (mL) *
 *nI-10uM*: Normalized intensity of A280 trace (unitless) ### Figure1D ####
 Fig1D_raw.csv * *frame_indices*: Indices of images containing binding
 events (unitless) * *x_coords*: x-coordinate of contrast signal *
 *y_coords*: y-coordinate of contrast signal * *contrasts*: Ratiometric
 contrast (intensity) * *calibrated_values*: Computed mass of each binding
 event (kDa) * *nearest_neighbour_distances*: Distance to nearest binding
 event (px), note: events under minimum allowable distance are removed from
 the histogram * *selections*: Number of events observed at this index and
 coordinate set ### Figure2A #### 20241209_10uMRho_IST100.txt * *Header*: 7
 lines summarizing acquisition parameters and the file structure * *Mass*:
 Mass-to-charge ratio of ions detected (m/z, Th) * *Intensity*: Measure of
 ion abundance (intensity units) #### 20241209_10uMRho-10uMATP_IST100.txt *
 *Header*: 7 lines summarizing acquisition parameters and the file
 structure * *Mass*: Mass-to-charge ratio of ions detected (m/z, Th) *
 *Intensity*: Measure of ion abundance (intensity units) ### Figure2B ####
 20241222_10uMRho-0uMATP_IST75.txt * *Header*: 7 lines summarizing
 acquisition parameters and the file structure * *Mass*: Mass-to-charge
 ratio of ions detected (m/z, Th) * *Intensity*: Measure of ion abundance
 (intensity units) #### 20241222_10uMRho-1uMATP_IST75.txt * *Header*: 7
 lines summarizing acquisition parameters and the file structure * *Mass*:
 Mass-to-charge ratio of ions detected (m/z, Th) * *Intensity*: Measure of
 ion abundance (intensity units) #### 20241222_10uMRho-3uMATP_IST75.txt *
 *Header*: 7 lines summarizing acquisition parameters and the file
 structure * *Mass*: Mass-to-charge ratio of ions detected (m/z, Th) *
 *Intensity*: Measure of ion abundance (intensity units) ####
 20241222_10uMRho-5uMATP_IST75.txt * *Header*: 7 lines summarizing
 acquisition parameters and the file structure * *Mass*: Mass-to-charge
 ratio of ions detected (m/z, Th) * *Intensity*: Measure of ion abundance
 (intensity units) #### 20241222_10uMRho-7uMATP_IST75.txt * *Header*: 7
 lines summarizing acquisition parameters and the file structure * *Mass*:
 Mass-to-charge ratio of ions detected (m/z, Th) * *Intensity*: Measure of
 ion abundance (intensity units) #### 20241222_10uMRho-20uMATP_IST75.txt *
 *Header*: 7 lines summarizing acquisition parameters and the file
 structure * *Mass*: Mass-to-charge ratio of ions detected (m/z, Th) *
 *Intensity*: Measure of ion abundance (intensity units) ####
 20241222_10uMRho-40uMATP_IST75.txt * *Header*: 7 lines summarizing
 acquisition parameters and the file structure * *Mass*: Mass-to-charge
 ratio of ions detected (m/z, Th) * *Intensity*: Measure of ion abundance
 (intensity units) #### 20241222_10uMRho-100uMATP_IST75.txt * *Header*: 7
 lines summarizing acquisition parameters and the file structure * *Mass*:
 Mass-to-charge ratio of ions detected (m/z, Th) * *Intensity*: Measure of
 ion abundance (intensity units) ### FigureS1 ####
 20240716_2uMRho_IST200_HCD200.txt * *Header*: 7 lines summarizing
 acquisition parameters and the file structure * *Mass*: Mass-to-charge
 ratio of ions detected (m/z, Th) * *Intensity*: Measure of ion abundance
 (intensity units) ### FigureS2A #### 20241209_10uMRho-emitter1_IST30.txt *
 *Header*: 7 lines summarizing acquisition parameters and the file
 structure * *Mass*: Mass-to-charge ratio of ions detected (m/z, Th) *
 *Intensity*: Measure of ion abundance (intensity units) ####
 20241209_10uMRho-emitter1_IST100.txt * *Header*: 7 lines summarizing
 acquisition parameters and the file structure * *Mass*: Mass-to-charge
 ratio of ions detected (m/z, Th) * *Intensity*: Measure of ion abundance
 (intensity units) #### 20241209_10uMRho-emitter1_IST200.txt * *Header*: 7
 lines summarizing acquisition parameters and the file structure * *Mass*:
 Mass-to-charge ratio of ions detected (m/z, Th) * *Intensity*: Measure of
 ion abundance (intensity units) #### 20241212_10uMRho-emitter2_IST200.txt
 * *Header*: 7 lines summarizing acquisition parameters and the file
 structure * *Mass*: Mass-to-charge ratio of ions detected (m/z, Th) *
 *Intensity*: Measure of ion abundance (intensity units) ### FigureS2B ####
 20250102_2uMRho-emitter1_IST75.txt * *Header*: 7 lines summarizing
 acquisition parameters and the file structure * *Mass*: Mass-to-charge
 ratio of ions detected (m/z, Th) * *Intensity*: Measure of ion abundance
 (intensity units) #### 20241222_2uMRho-emitter2_IST75.txt * *Header*: 7
 lines summarizing acquisition parameters and the file structure * *Mass*:
 Mass-to-charge ratio of ions detected (m/z, Th) * *Intensity*: Measure of
 ion abundance (intensity units) #### 20250102_2uMRho-emitter1_IST200.txt *
 *Header*: 7 lines summarizing acquisition parameters and the file
 structure * *Mass*: Mass-to-charge ratio of ions detected (m/z, Th) *
 *Intensity*: Measure of ion abundance (intensity units) ####
 20241222_2uMRho-emitter2_IST200.txt * *Header*: 7 lines summarizing
 acquisition parameters and the file structure * *Mass*: Mass-to-charge
 ratio of ions detected (m/z, Th) * *Intensity*: Measure of ion abundance
 (intensity units) ### FigureS3 #### 20240908_S200Increase_Calibration.csv
 * *running_volume*: Volume of running buffer passed through the column
 (mL) * *mAU*: Absorbance at A280 (mAU) * *mass_standards*: Names of
 species used to calibrate the column (categorical) * *Daltons*: Molecular
 weights of species used to calibrate the column (Da) * *elution_volume*:
 Volume at apex of peak assigned to each calibrant (mL) * *log(Daltons)*:
 Exponent of molecular weight in base 10 (unitless) * *Kav*: Partition
 coefficient to represent the separation range of the column (unitless) *
 *column_volume*: Volume of column bed, used to calculate the partition
 coefficient (mL) * *void_volume*: The volume where the largest,
 unseparated components of the injection begin to elute from the column,
 used to calculate the partition coefficient (mL) ### FigureS4 ####
 FigS4_raw.csv * *frame_indices*: Indices of images containing binding
 events (unitless) * *x_coords*: x-coordinate of contrast signal *
 *y_coords*: y-coordinate of contrast signal * *contrasts*: Ratiometric
 contrast (intensity) * *calibrated_values*: Computed mass of each binding
 event (kDa) * *nearest_neighbour_distances*: Distance to nearest binding
 event (px), note: events under minimum allowable distance are removed from
 the histogram * *selections*: Number of events observed at this index and
 coordinate set ### FigureS5 #### MP_oligomer_populations.csv * *micromolar
 ATP*: Concentration of ATP in micromolar (uM) * *monomer*: Percentage of
 particles falling under the monomer distribution (%) * *stdev-m*: Standard
 deviation of monomer replicates (%) * *dimer*: Percentage of particles
 falling under the dimer distribution (%) * *stdev-d*: Standard deviation
 of dimer replicates (%) * *trimer*: Percentage of particles falling under
 the trimer distribution (%) * *stdev-t*: Standard deviation of trimer
 replicates (%) * *tetramer*: Percentage of particles falling under the
 tetramer distribution (%) * *stdev-q*: Standard deviation of tetramer
 replicates (%) * *pentamer*: Percentage of particles falling under the
 pentamer distribution (%) * *stdev-p*: Standard deviation of pentamer
 replicates (%) * *hexamer*: Percentage of particles falling under the
 hexamer distribution (%) * *stdev-h*: Standard deviation of hexamer
 replicates (%) ### FigureS6 #### 20241209_10uMRho-10uMATP_IST30.txt *
 *Header*: 7 lines summarizing acquisition parameters and the file
 structure * *Mass*: Mass-to-charge ratio of ions detected (m/z, Th) *
 *Intensity*: Measure of ion abundance (intensity units) ####
 20241209_10uMRho-10uMATP_IST50.txt * *Header*: 7 lines summarizing
 acquisition parameters and the file structure * *Mass*: Mass-to-charge
 ratio of ions detected (m/z, Th) * *Intensity*: Measure of ion abundance
 (intensity units) #### 20241209_10uMRho-10uMATP_IST100.txt * *Header*: 7
 lines summarizing acquisition parameters and the file structure * *Mass*:
 Mass-to-charge ratio of ions detected (m/z, Th) * *Intensity*: Measure of
 ion abundance (intensity units) #### 20241209_10uMRho-10uMATP_IST150.txt *
 *Header*: 7 lines summarizing acquisition parameters and the file
 structure * *Mass*: Mass-to-charge ratio of ions detected (m/z, Th) *
 *Intensity*: Measure of ion abundance (intensity units) ####
 20241209_10uMRho-10uMATP_IST200.txt * *Header*: 7 lines summarizing
 acquisition parameters and the file structure * *Mass*: Mass-to-charge
 ratio of ions detected (m/z, Th) * *Intensity*: Measure of ion abundance
 (intensity units) ### FigureS7 #### 20241212_10uMRho-100uMATP_IST75 *
 *Header*: 7 lines summarizing acquisition parameters and the file
 structure * *Mass*: Mass-to-charge ratio of ions detected (m/z, Th) *
 *Intensity*: Measure of ion abundance (intensity units) ### FigureS8 ####
 20241222_10uMRho-0uMATP_IST75.txt * *Header*: 7 lines summarizing
 acquisition parameters and the file structure * *Mass*: Mass-to-charge
 ratio of ions detected (m/z, Th) * *Intensity*: Measure of ion abundance
 (intensity units) #### 20241222_10uMRho-1uMATP_IST75.txt * *Header*: 7
 lines summarizing acquisition parameters and the file structure * *Mass*:
 Mass-to-charge ratio of ions detected (m/z, Th) * *Intensity*: Measure of
 ion abundance (intensity units) #### 20241222_10uMRho-3uMATP_IST75.txt *
 *Header*: 7 lines summarizing acquisition parameters and the file
 structure * *Mass*: Mass-to-charge ratio of ions detected (m/z, Th) *
 *Intensity*: Measure of ion abundance (intensity units) ####
 20241222_10uMRho-5uMATP_IST75.txt * *Header*: 7 lines summarizing
 acquisition parameters and the file structure * *Mass*: Mass-to-charge
 ratio of ions detected (m/z, Th) * *Intensity*: Measure of ion abundance
 (intensity units) #### 20241222_10uMRho-7uMATP_IST75.txt * *Header*: 7
 lines summarizing acquisition parameters and the file structure * *Mass*:
 Mass-to-charge ratio of ions detected (m/z, Th) * *Intensity*: Measure of
 ion abundance (intensity units) #### 20241222_10uMRho-20uMATP_IST75.txt *
 *Header*: 7 lines summarizing acquisition parameters and the file
 structure * *Mass*: Mass-to-charge ratio of ions detected (m/z, Th) *
 *Intensity*: Measure of ion abundance (intensity units) ####
 20241222_10uMRho-40uMATP_IST75.txt * *Header*: 7 lines summarizing
 acquisition parameters and the file structure * *Mass*: Mass-to-charge
 ratio of ions detected (m/z, Th) * *Intensity*: Measure of ion abundance
 (intensity units) #### 20241222_10uMRho-100uMATP_IST75.txt * *Header*: 7
 lines summarizing acquisition parameters and the file structure * *Mass*:
 Mass-to-charge ratio of ions detected (m/z, Th) * *Intensity*: Measure of
 ion abundance (intensity units) #### rho_fitting_example.py *
 *Description*: Script used to fit ATP-bound populations of +32 charge
 state of the Rho hexamer. Includes an example to generate synthetic
 experiments (lines 10-17). Initial parameters are provided to
 `itc_model.set_params` in units of kilojoules per mole.