First pore and tunnel analyses

Use the bundled five-model gramicidin A structure to verify the installation and produce a pore profile, then use the bundled cholesterol oxidase structure to find tunnels from internal cavities.

Pore mode

1. Load the structure

In VMD, select File → New Molecule…, browse to vmdhole/1GRM.pdb, and select Load. Then open Extensions → Analysis → VMDHole and select Pore.

2. Set the input

Set the following controls:

Control	Value
Molecule	the loaded 1GRM molecule
Selection	all
Frames	now

The all selection includes gramicidin's nonstandard terminal residues. For another system, start with protein and add any cofactor, ligand, or blocker that forms part of the pore wall. Exclude bulk water, membrane atoms, and freely moving ions unless they are intentionally part of the obstruction.

Open File → Settings and confirm that Radius file points to a readable HOLE .rad file. Every selected atom, including nonstandard residues and cofactors, must match a radius rule. Do not use a result if the VMD console reports missing radii; choose or edit an appropriate radius file first. See Radius files.

3. Check the pore axis

CPOINT should lie inside the channel, and CVECT should point along it. When a molecule is activated, VMDHole proposes suitable values from the current selection and structure. Keep them for this first run if the point and direction look correct.

Open the HOLE parameters gear and enable Show cues. Confirm that the CPOINT marker is inside the channel and the CVECT arrow follows its long axis.

To define the gramicidin axis explicitly:

1. Select ⌖ beside CVECT.
2. Enter resname ETA and chain A for Point 1.
3. Enter resname ETA and chain B for Point 2.
4. Select Compute and inspect the displayed direction.

The vector sign changes the profile orientation, not the pore being analysed. For another structure, use Guess, Use Z, two selections marking the pore ends, or two coordinates.

4. Run the current conformation

Keep the initial spherical-method defaults and select Run HOLE. A completed run should show:

• a radius profile in Pore Profile;
• the pore centreline or surface in the VMD display;
• the minimum-radius readout for the current frame.

Inspect the 3D result before using the numbers. The centreline must pass through the intended channel rather than an external groove. If it does not, correct CPOINT, CVECT, or Selection, then rerun.

5. Compare all five conformers

Set Frames to all and select Run HOLE again. Use:

• Over Time to compare position across conformers;
• Mean Profile to inspect the mean and spread;
• Trends to compare minimum radius, volume, or conductance estimates;
• Histogram to inspect the distribution of per-frame radii.

These five experimentally determined NMR conformers show structural variation and the corresponding variation in pore geometry. Unlike consecutive MD frames, however, their order does not encode elapsed time.

Tunnel mode

1. Load the structure

Load vmdhole/1MXT.pdb in VMD and select Tunnel in VMDHole. Set:

Control	Value
Molecule	the loaded 1MXT molecule
Selection	protein
Frames	now

2. Find and inspect tunnels

Enter 20.4632 0.4374 17.4692 under Start point. Open the MOLE parameters gear, enable Show cues, confirm that the marker lies inside the protein, and select Run Tunnel.

The route table lists the tunnels found from qualifying internal cavities. Select a route and confirm in the VMD display that it begins inside the structure and reaches the molecular surface. Use Lining… to inspect its contacting residues. Route rank is a search score; visual inspection remains part of validating a candidate pathway.

Next steps

Choose the Pore or Tunnel tutorial for the mode you just ran. Afterward, use the matching workflow guide when you need more than the tutorial settings.