Dataset DOI: 10.5061/dryad.44j0zpcvz

Description of the data and file structure

The data falls into 3 broad categories:

Optical Trapping

TIRF Microscopy

Refeyn

Refeyn:

The raw Refeyn data was taken on a TwoMP mass photometer, and the calibrated files has been exported in the .csv format.

TIRF:

The TIRF microscopy data covers three experiments. In the first, oligo-labeled FL KIF5B is mixed with a complementary fluorescent DNA oligo and allowed to walk on microtubules decorated with MAP7. The 488 channel is KIF5B-GFP, and the other channel is the Cy3 oligo. This data was used exclusively for illustrative purposes and was not analyzed.

In the second, FL KIF5B is attached to streptavidin quantum dots via a DNA handle and allowed to walk on axonemes decorated with MAP7. This assay was used as a control for the DNA handle attachment system and to measure the velocity and run length of KIF5B on axonemes.

Lastly, k560-GFP was allowed to walk on axonemes in the absence of MAP7. These movies were used only to measure the velocity and run length of k560 on axonemes.

Optical Trapping:

Two types of optical trapping experiments were performed.

In one type, motors were allowed to walk against a fixed trap. These experiments were performed for k560 attached directly to a bead, k560 attached to a bead via a DNA tether, FL KIF5B attached to a bead via a DNA tether, and chassis experiments where two or three copies of k560 were attached to a DNA chassis which was then either directly attached to the bead or via a DNA tether. This data is presented as .csv files where one column each corresponds to time, force parallel to the axoneme, trap position perpendicular to the axoneme, force perpendicular to the axoneme, and trap position parallel to the axoneme.

In the other type, the trap position was updated to provide a fixed load. These experiments were performed for k560 attached directly to beads and FL KIF5B attached via a DNA tether. This data is presented as .csv files, where one column each corresponds to time, displacement along the axoneme, trap position perpendicular to the axoneme (always zero), displacement perpendicular to the axoneme, and trap position along the axoneme.

Files and variables

File: Raw_Data_for_Z-Force_Project.zip

Description: Raw data for entire publication

Fixed Trap: All files are called 'TraceX.csv' and contain time in seconds (first column), force along the axis of the axoneme in pN (column 2), trap position along the axis perpendicular to the axoneme in nm (column 3), force along the axis perpendicular to the axoneme in pN (column 4), and trap position along the axis parallel to the axoneme in nm (column 5). Since this is a fixed trap assay, the values in columns 3 and 5 are always zero.

-->Chassis

-->-->High Z-Force

2 Binding Sites

3 Binding Sites

-->-->Low Z-Force

1 Binding Site

2 Binding Sites

3 Binding Sites

-->Single Motor

-->-->High Z-Force

-->-->Low Z-Force

Force Feedback: All files are called 'TraceX.csv' and contain time in seconds (first column), displacement along the axis of the axoneme in nm (column 2), trap position along the axis perpendicular to the axoneme in nm (column 3), displacement along the axis perpendicular to the axoneme in nm(column 4), and trap position along the axis parallel to the axoneme in nm (column 5). Since force feedback is only performed on the axis of the axoneme, the values in column 3 are always zero.

-->High Z-Force

+0.5 pN

+2 pN

+3 pN

+4 pN

-1 pN

-2 pN

-4 pN

-6 pN

-10 pN

-->Low Z-Force

+0.5 pN

+2 pN

+3 pN

+4 pN

-1 pN

-2 pN

-4 pN

-6 pN

-10 pN

Refeyn: All files are .csv format. The first column is the frame. Note that most frames have more than one particle. Columns 2 and 3 are the x- and y- coordinates of a particle, respectively, sub-pixel localized from a Gaussian fit of the interference pattern. Column 4 is the change in light intensity at the particle (unitless ratio). The fifth column is the fitted molecular weight in kD. The sixth column is the fit error in kD. The seventh column is error related to a differential blurring filter, which was not used. As such, the seventh column is uniformly nan. The eighth column is nearest neighbor distance in pixels. The ninth column is a Boolean for whether or not the particle is chosen for analysis, which always has a value of 1.

-->High Z - 2 Binding Sites

-->High Z - 3 Binding Sites

-->Kinesin Alone

-->Low Z - 2 Binding Sites

-->Low Z - 3 Binding Sites

TIRF: All files are in .tif format.

-->FL KIF5B

-->-->Handle Control w QDots on Axonemes (pixel size 43 nm)

-->-->Oligo Label Control w Complementary Oligo on MTs (pixel size 160 nm)

-->K560

-->-->Motility on Axonemes (pixel size 160 nm)

Code/software

ImageJ is commonly used to analyze .tif files.

Users may find custom MATLAB software helpful for analyzing force feedback measurements (https://github.com/Yildiz-Lab/ForceFeedbackAutoAnalyzer).

Access information

Other publicly accessible locations of the data:

• 
  

Data was derived from the following sources:

•