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Step 6: Calculation of dissociation free energy (HPC/local)

In the previous Step 5 , we have generated in each of the 384 bound folders (bound/fsdam/bi, with 1 < i < 384) and in each of the 192 the unbound folders (unbound/fsdam/ui, with 1< i <192) two dhdl.xvg files, namely dhdlQ.xvg and dhdlvdw.xvg, relative to electrostatic and LJ ligand-environment interactions, respectively.
These files are processed to obtain the absolute dissociation free energy using the script works.bash which returns 4 estimates for the ligand-receptor dissociation free energy: 3 estimates are based on Gaussian mixtures (one, two and three components) and the Jarzynski estimate.

Untar the bin archive in the parent directory (USER_SCRATCH) where you copied the bound and unbound dir directories (see Step 3 ), and do:

	cd USER_SCRATCH
	cd bin/em
	make
	

Then, jump back to the parent directory USER_SCRATCH and launch from this directory the works.bash script as

	bin/works.bash PF-3clpro 
     

PF-3clpro is the user-defined name for the project. The output of the program should be the following:

lj bound done
qq bound done
lj unbound done
qq unbound done
 DG1= 2.0  2.2 DG2=  1.5  4.5 DG3=  3.9  4.7 DGj=  8.6  0.9

The last line refer to the four estimates of the dissociation free energy (in kcal/mol) for the complex PF-07321332-3CL^pro (the second number is the 95% confidence interval). The first three estimates, DG1, DG2, DG3 have been computed using the em program (based on the Expectation Maximization algorithm) where the bound and unbound vDSSB convolution work distribution have been fitted using Gaussian mixtures with 1 to 3 components. The large confidence interval for these EM-based estimate denotes poor fitting suggesting that the convolution work distribution is likely to be due to >3 Gaussian components. The last numbers (8.6 0.9) refers to the Jarzynski estimate, which is asymptotically exact and is taken as the more reliable estimate in the present case. The script generates in a subdirectory Results a series of files with the basename PF-3clpro containing the salient processed data for the PF-07321332-3CL^pro example.

To calculate the volume correction to dissociation free energy, the script VOLcor.bash can be used. It acts on the pullx.xvg files (see Zenodo repository) generated in the target state replica of each BATTERY of the bound state, saving the correction to the Results directory. For the PF-07321332-3CL^pro complex the correction should be equal to -2.8 kcal/mol. In the SI.zip attached to the JCIM note we also include the script Qcor.sh for the finite-size correction applying to charged ligands. Such correction dose not apply to PF-07321332-3CL^pro as the ligand bears no net charge. For further technical details on the usage of these scripts see the README files in the 6_post directory of the SI.zip archive attached to the JCIM note.