X-ray data processing

Prepare data#

Convert h5 format to cbf format: XDS can't operate with h5 format images, so you need to convert the image to cbf format before the process. On macOS, eiger2cbf is a useful program to do this.

eiger2cbf path/to/your/h5_master_data           -- get number of frames
eiger2cbf path/to/your/h5_master_data N out.cbf -- write N-th frame to out.cbf
eiger2cbf path/to/your/h5_master_data N         -- write N-th frame to STDOUT
eiger2cbf path/to/your/h5_master_data N:M   out -- write N to M-th frames to outNNNNNN.cbf

For example:

eiger2cbf path/to/your/h5_master_data 1:720 out # convert all the 720 images to cbf format

X-Ray data processing#

Process with XDS#

XDSGUI

Install XDSGUI according to this instrucment.
Run the XDSGUI in the terminal.
```
cd path/to/your/images
xdsgui
```
Load the image and generate the XDS.INP file:
Modify the XDS.INP file to meet your needs, then click Run XDS.

If you collect your data with 10U2 beamline in SSRF, you should modify the ROTATION_AXIS parameter to 0 -1 0.
Check the result in page CORRECT

Process with Xia2#

There are two ways to use Xia2: use Xia2 in CCP4, or install the latest xia2/DIALS bundle according to this website.

Process your data with it:

cd path/to/images
xia2 pipeline=dials-aimless ./ goniometer.axes=0.000000,1.000000,0.000000 xia2.settings.resolution.d_min=2

Process with autoPROC#

Request a license and install autoPORC according to this website.

Process your data with it:

cd path/to/images
process -I . -d autoproc -R 50 2 > out.log 
#save the result in subdirectory named autoproc, resolution vary from 2 to 50, save the log to out.log file.

Structure determination#

Valid the data#

Find the XDS_ASCII.HKL file in the autoproc directory as the diffraction file.
Valid the data with Phenix Xtriage, put the diffraction file in, and run.

Molecular replacement#

Find the XDS_ASCII.HKL file in the autoproc directory as the diffraction file.
Get the template model:
1. Blast the sequence of your protein with the PDB database, select the proper structure, and download the PDB file.
1. After downloading the template model, you need to edit it with PyMOL to remove the redundant copies and water molecules.
Determine how many copies are in ASU(asymmetric unit):
1. You can get the number from the result of the Xtriage
  
  ::: grid {cols=2,rows=1,gap=12,type=images}
  
  :::
2. Or you can use the cell content analysis tool in CCP4.
[!NOTE]

You can determine the number of copies through the Solvent content, generally, the solvent content of most biological macromolecule crystals is between 40% and 60%.
Do molecular replacement with Phenix Phaser-MR.

::: grid {cols=4,rows=1,gap=12,type=images}

:::
Build your model with Phenix AutoBuild: input the result of molecular replacement, and run.
Check the model from AutoBuild in Coot and refine it manually.

::: grid {cols=2,rows=1,gap=12,type=images}

:::

此文由 Mix Space 同步更新至 xLog
原始链接为 https://xxu.do/posts/academic/X-ray-data-processing