Structure calculation

Step 1. Load Data.

 

• • open new project PRJ3 [Project>new]
  • load 15N NOESY peak list [DATA>N15 NOESY>load]
  • load 13C_aliphatic NOESY peak list [DATA>C13NOESY H2O>load]
  • load 13C_aromatic NOESY peak list [DATA>AromNOESY>load]
  • set tolerances [Data>Tolerances]
  • load 13C NOESY peak list [DATA>C13NOESY H2O>load]
  • load assigned chemical shifts [Fragment>Load>assigned]

            The file with assigned chemical shifts could be either in “standard” format   (assigned AA-fragments) or in cyana format (prot-file).

 

     

Step 2. Set up constraints

 

 The structure calculation requires dihedral angle constraints in the cyana format (aco-file). These constraints are usually prepared using the results of dihedral angle prediction by TALOS. H-bond constraints are optional.

 

-          calculate dihedral angle constraints [Structure>Constraints>Talos>Calculate] 

-          set up H-bond constraints [Structure>Constraints>H-bonds>Specify] 

           

In the cased dihedral angle or H-bond constraints in cyana format (aco-file or upl-file, respectively) already are  prepared,  then the constraints can be loaded  from the corresponding files   [Structure->Constraints->Talos>Load] or [Structure>Constraints>H-bonds>Load]

 

 

Step 3. Specify ligands coordinating ZN ions (if there are any).

 

If there are ZN ions as a part of a protein structure the file “zn_ligands” should be present inside FMCGUI project directory. This file can be created by the following command

 

      - specify residues that coordinate ZN ion(s) [Structure>Add ZN ion]

 

 

Step 4. Set up CYANA calculations

 

       - setting up structure calculations with CYANA [Structure>Calcuate>cyana]

          All files that are necessary for CYANA run are prepared and saved in the user specified directory, crun#, which is located inside the project directory. These files include chemical shifts (belok.prot file), sequence file, peak lists, dihedral angles constraints (file belok.aco), H-bond constraints, if available, (file hbond.upl), and constraints for ZN ions, if present, (files zn.upl, zn.lol).

 

Step 5. Structure evaluation and peak list refinement.

 

RPF analysis and DP score allow one to estimate goodness-of-fit of a structural ensemble to NOESY peak lists. The results of RPF analysis can serve both for structure validation and peak lists refinement.

 

-          run RPF analysis [Structure>RPF>RPThe results of the RPF analysis include peak lists in the SPARKY format of both false negative and false positive peaks for C13_aliphatic_NOESY, C13_aromatic_NOESY, and N15_NOESY spectra in separate files.].

-          set up DP-score calculations with AutoStructure [Structure>RPF>DP].