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Alanine scanning

This example estimates the effect of mutating receptor residue Tyr23 to alanine in a protein-protein complex. It uses the same solvated simulation, frame range, and GB-Neck2 model as the decomposition and stability examples.

  • Analysis

    Computational alanine scanning

  • Mutation

    R:23 Tyr to Ala

  • Solvent model

    GB-Neck2 (igb=8)

  • Bundled test

    gmx_MMPBSA_test -t 12

Before you begin

The manual workflow uses the following files and selections:

  • Calculation settings

    mmpbsa.in (-i)

  • GROMACS system

    Structure com.tpr (-cs) and topology topol.top (-cp). Keep the toppar directory containing the referenced *.itp files beside topol.top.

  • Trajectory

    PBC-corrected and fitted trajectory com_traj.xtc (-ct)

  • Molecular selections

    Index index.ndx (-ci) with the SOLU_chain1 and SOLU_chain2 groups (-cg)

The folder includes the solvated system snapshot com.pdb, matching the corresponding files in the decomposition and stability examples. The 608-atom reference.pdb structure (-cr) preserves the chain ID and residue number used by mutant_res="R:23". See the complete command-line reference for all options.

Run the example

Run the bundled test

The quickest way to reproduce this example is through the test runner:

gmx_MMPBSA_test -t 12

See the gmx_MMPBSA_test documentation for download, selection, and cleanup options.

Run it manually

Download the Alanine Scanning example as a ZIP archive.

Extract the archive, change to the Alanine_scanning directory, and choose either the serial or MPI command. You can also view the example files on GitHub before downloading them.

gmx_MMPBSA -O \
  -i mmpbsa.in \
  -cs com.tpr \
  -ct com_traj.xtc \
  -ci index.ndx \
  -cg SOLU_chain1 SOLU_chain2 \
  -cp topol.top \
  -cr reference.pdb \
  -o FINAL_RESULTS_MMPBSA.dat \
  -eo FINAL_RESULTS_MMPBSA.csv

mpirun -np 2 gmx_MMPBSA -O \
  -i mmpbsa.in \
  -cs com.tpr \
  -ct com_traj.xtc \
  -ci index.ndx \
  -cg SOLU_chain1 SOLU_chain2 \
  -cp topol.top \
  -cr reference.pdb \
  -o FINAL_RESULTS_MMPBSA.dat \
  -eo FINAL_RESULTS_MMPBSA.csv

Configure the calculation

The example uses the minimal mmpbsa.in shown first below. The all-options version was generated with gmx_MMPBSA --create_input gb ala and then adapted with the same example-specific values. The concise block is the runnable starting point; the generated block includes additional options and defaults, so the two blocks are not textually identical. Both blocks therefore describe the same MM/GBSA alanine-scanning calculation.

mmpbsa.in
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Sample input file for Alanine scanning
# This input provides a practical starting point for alanine scanning.
# Verify the mutation selection and model settings for your system.

&general
sys_name="Alanine_Scanning",
startframe=1,
endframe=10,
PBRadii=4,
/
&gb
igb=8, saltcon=0.150,
/
&alanine_scanning
mutant='ALA', mutant_res='R:23', cas_intdiel=1,
/
mmpbsa.in generated with --create_input gb ala
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Input block generated for the 1.7.0 release.
Be careful with the variables you modify, some can have severe consequences on the results you obtain.

# General namelist variables
&general
  sys_name                       = "Alanine_Scanning"                       # System name; e.g. "complex"
  startframe                     = 1                                      # First frame; e.g. 1
  endframe                       = 10                                     # Last frame; e.g. 100
  interval                       = 1                                      # Frame interval; e.g. 1


  PBRadii                        = 4                                      # PB radii set; 1-7
  temperature                    = 298.15                                 # Temperature (K); e.g. 298.15
  qh_entropy                     = 0                                      # Legacy QH output reader; new calculations reject 1
  interaction_entropy            = 0                                      # Run IE entropy; 0/1
  ie_segment                     = 25                                     # IE tail diagnostic only (%); not primary IE; e.g. 25
  c2_entropy                     = 0                                      # Run C2 entropy; 0/1
  assign_chainID                 = 0                                      # Assign chain IDs; 0/1
  exp_ki                         = 0.0                                    # Experimental Ki (nM); e.g. 0.0
  full_traj                      = 0                                      # Write full trajectory; 0/1
  gmx_path                       = ""                                     # GROMACS path; e.g. "/usr/bin"
  keep_files                     = 2                                      # Files to keep; 0-2
  netcdf                         = 0                                      # Use NetCDF; 0/1
  solvated_trajectory            = 1                                      # Clean solvated traj.; 0/1
  explicit_waters                = 0                                      # Explicit waters; e.g. 10
  explicit_waters_mask           = "dASA"                                     # Water reference; e.g. ":1-10", "within 4", "dASA"
  explicit_waters_group          = "automatic"                                     # Solvent group; e.g. "TIP3" or "automatic"
  explicit_waters_dasa_cutoff    = 0.5                                    # dASA cutoff; e.g. 0.5
  explicit_waters_as             = "receptor"                             # Water owner; e.g. "receptor"
  explicit_waters_extra_points   = "error"                                # Virtual sites; "error" or "strip"
  verbose                        = 1                                      # Output verbosity; 0-2
/

# (AMBER) Generalized-Born namelist variables
&gb
  igb                            = 8                                      # GB model, e.g. 2 or 8
  intdiel                        = 1.0                                    # Internal dielectric; e.g. 1.0
  extdiel                        = 78.5                                   # External dielectric; e.g. 78.5
  saltcon                        = 0.150                                  # Salt conc. (M); e.g. 0.150
  surften                        = 0.0072                                 # Surface tension; e.g. 0.0072
  surfoff                        = 0.0                                    # Surface offset; e.g. 0.0
  molsurf                        = 0                                      # Use molsurf; 0/1
  msoffset                       = 0.0                                    # Molsurf offset; e.g. 0.0
  probe                          = 1.4                                    # Probe radius (A); e.g. 1.4
  ifqnt                          = 0                                      # Enable QM/MM; 0/1
  qm_theory                      = "PM6-DH+"                              # QM theory; e.g. "PM6-DH+"
  qm_residues                    = ""                                     # QM residues; e.g. ":1-5"
  com_qmmask                     = ""                                     # Complex QM mask; e.g. ":1-5"
  rec_qmmask                     = ""                                     # Receptor QM mask; e.g. ":1-5"
  lig_qmmask                     = ""                                     # Ligand QM mask; e.g. ":1"
  qmcharge_com                   = 0                                      # Complex QM charge; e.g. 0
  qmcharge_lig                   = 0                                      # Ligand QM charge; e.g. 0
  qmcharge_rec                   = 0                                      # Receptor QM charge; e.g. 0
  qmcut                          = 9999.0                                 # QM cutoff (A); e.g. 9999
  scfconv                        = 1e-08                                  # SCF convergence; e.g. 1.0e-8
  itrmax                         = 1000                                   # Maximum SCF iterations; e.g. 5000
  # ndiis_attempts                 = None                                 # Maximum DIIS attempts per SCF cycle; e.g. 700
  peptide_corr                   = 0                                      # Peptide correction; 0/1
  writepdb                       = 1                                      # Write QM PDB; 0/1
  verbosity                      = 0                                      # QM/MM verbosity; 0-5
  alpb                           = 0                                      # Use ALPB; 0/1
  arad_method                    = 1                                      # ALPB size method; e.g. 1
/

# Alanine scanning namelist variables
&alanine_scanning
  mutant_res                     = "R:23"                                     # Residue to mutate; e.g. "A/23"
  mutant                         = "ALA"                                  # Mutation target; "ALA" or "GLY"
  mutant_only                    = 0                                      # Mutant energies only; 0/1
  cas_intdiel                    = 1                                      # Set intdiel by residue; 0/1
  intdiel_nonpolar               = 1                                      # Nonpolar intdiel; e.g. 1
  intdiel_polar                  = 3                                      # Polar intdiel; e.g. 3
  intdiel_positive               = 5                                      # Positive intdiel; e.g. 5
  intdiel_negative               = 5                                      # Negative intdiel; e.g. 5
/

Keep in mind

This input provides a practical starting point and can serve as the basis for production calculations. Review the available input-file options, their accepted values, and adjust settings that depend on your system or protocol. Additional sample inputs are available here.

How this example works

The single-trajectory calculation selects the 608-atom protein-protein solute from the solvated source trajectory. It processes frames 1 through 10 with GB-Neck2 (igb=8), the matching mbondi3 radii (PBRadii=4), and a salt concentration of 0.15 M.

mutant_res="R:23" identifies Tyr23 in receptor chain R, and mutant="ALA" creates the corresponding alanine variant without requiring a separately prepared mutant topology. With mutant_only=0, the calculation evaluates both the original and mutant systems. Because tyrosine is classified as polar, cas_intdiel=1 applies intdiel_polar=3 to the relevant GB calculations.

Verify the mutation selection

Residue selection depends on stable chain IDs and residue numbers. Supplying reference.pdb with -cr makes the mapping explicit while com.pdb retains the complete solvated snapshot for consistency. Confirm the selected mutation in _GMXMMPBSA_COM_FIXED.pdb and the generated mutant structures before interpreting results.

One run can mutate one residue or several residues together in one composite mutant; it does not independently scan each selected residue. mutant_res also accepts CHAIN/RESNUM notation and insertion codes as described in the alanine-scanning input options.

Expected outputs

A successful calculation produces:

  • FINAL_RESULTS_MMPBSA.dat: the original, mutant, and alanine-scanning energy summaries.
  • FINAL_RESULTS_MMPBSA.csv: the per-frame energy terms requested with -eo.

Analyze the results

Open the results with gmx_MMPBSA_ana for interactive inspection and plotting. See the gmx_MMPBSA_ana documentation for usage details.

Last update: September 13, 2026 05:57:32
Created: February 8, 2021 07:10:13
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