Par file in refinement mode

Hi Matt,

Sorry to hear you're having trouble.

The INPUT_PAR_FILENAME should point to the waypoints.ali file from the startup run.

I would recommend trying out the flx_wrap.rb script to run Frealix, since this will handle this level of detail for you. Have you tried it yet?

For example, in your case, create a folder named frealix_round_000 and make sure it contains a waypoints.ali file (a copy of the output file from the startup run), as well as your initial model 3D reconstruction (a copy of the 3drec* files from the startup run). Then run something like:
flx_wrap.rb -r 1 -s 20.0 -f 20.0 your_parameter_file.txt

where your_parameter_file is a barebones set of parameters, which should contain (at a minimum) the following keys:

• ctffind_files or ctftilt_files (give "none" for the one you won't use)
• film_files
• twist_per_subunit
• rise_per_subunit
• pixel_size
• helix_max_radius

Here is an example of such a file from one of my amyloid projects:

    FILM_SELECTION                  1-589
    FILM_FILES                      /my/folder/of/micrographs/movies_allframes_nofilter_magdistcorrectlinear.mrc

    CTFFIND_FILES                   /my/folder/of/ctf/movies_allframes_nofilter_magdistcorrectlinear_bin2_ctf.txt
    CTFTILT_FILES                   none

    TWIST_PER_SUBUNIT               1.54    #0.769  #degrees
    RISE_PER_SUBUNIT                9.4     #4.7    #in Angstroms
    SYMMETRY_AXIAL                  1
    SYMMETRY_PERP                   1
    FAL_WPS_IN_SCORE                T       # Whether the first and last segments of the filament should be included in the scoring function
    FAL_WPS_IN_THREED               F       # Whether the first and last segments of the filament should be included in the 3D reconstruction
    HELIX_MAX_RADIUS                80      #in Angstroms
    ASYM_UNIT_MW                    17672.4 # 4418.1 #MW for the 26-residue fragment used for synth #4418.1 for Abeta(1-42) #4329.9 is the MW for Abeta(1-40)
    STACK                           T

    PIXEL_SIZE                      1.04

    POLARITY_CHECK                  F
    PHI_POLARITY_CHECK              F
    REFINE_WPS_SIMULTANEOUSLY       F

    REFINE_X                        T
    REFINE_Y                        T
    REFINE_Z                        F
    REFINE_PHI                      T
    REFINE_THETA                    F
    REFINE_PSI                      T
    REFINE_SUBUNIT_NUM              F

    FRAGMENTED_SUBUNIT_NUM          T
    CONSTANT_RISE_PER_SUBUNIT       T
    CONSTANT_TWIST_PER_SUBUNIT      F
    AMYLOID_BETA                    T

    FORCE_WP_TO_WP_PHI              90.0

    SINGLE_FILAMENT_RECONSTRUCTIONS T       # output central slices of individual filament reconstructions to disk

    SEARCH_HALF_RANGE_X             1.0
    SEARCH_HALF_RANGE_Y             1.0
    SEARCH_HALF_RANGE_PSI           5.0
    SEARCH_HALF_RANGE_PHI           10.0
    SEARCH_HALF_RANGE_THETA         6.0

    SEARCH_STEP_SIZE_THETA          2
    SEARCH_FILAMENT_TILT            T

    BOOST_LOW_FREQUENCY_CONTRAST    T

    PSF_RADIUS                      200

Note that only a few of these parameters are necessary, but that you can add any of the parameters listed at the top of the output from mode filament_refine. In that output, you will also see a brief description of what each parameter does.

Also, film_files and ctffind_files can have wildcards in them, in which case the files need to be numbered consecutively with the numbers matching those in the "film" column of the waypoints.ali file.

Any parameter which you do not specify in the barebones parameter file should take up a sensible default value.

I hope this helps get you started - I apologise for the lack of documentation right now, but please don't hesitate to ask as many questions as it takes!

Alexis