Day 13: Macromolecular 3D Structure Exploration: PDB, Mol* & PyMOL Visualization

1. Macromolecular Structure Determination & PDB Coordinate Geometry

The 3D spatial conformation of a macromolecule determines its enzymatic activity, binding affinities, and therapeutic druggability. The Protein Data Bank (PDB) archives atomic coordinate files determined primarily through three experimental techniques:

2. Anatomy of the Legacy PDB and Modern mmCIF File Formats

Historically, structures were stored in the 80-column fixed-width PDB format. Because legacy PDB files cannot accommodate structures with > 62 chains or > 99,999 atoms (such as the 70S ribosome), the wwPDB transitioned to the mmCIF (macromolecular Crystallographic Information Framework) format:

# Legacy PDB ATOM record format:
# Columns:  1-6: Record name ("ATOM  ")
#           7-11: Atom serial number
#          13-16: Atom name (e.g. "CA", "N", "CB")
#          18-20: Residue name (e.g. "MET", "GLY")
#             22: Chain identifier (e.g. "A")
#          23-26: Residue sequence number
#          31-38: Orthogonal X coordinate in Angstroms
#          39-46: Orthogonal Y coordinate in Angstroms
#          47-54: Orthogonal Z coordinate in Angstroms
#          55-60: Occupancy (0.00 to 1.00)
#          61-66: Temperature factor / B-factor (isotropic thermal displacement)
ATOM    145  CA  ARG A  24     -10.421  24.512  14.288  1.00 18.45           C

3. Validation: Ramachandran Plots & Backbone Dihedral Angles

In 1963, G. N. Ramachandran demonstrated that steric clashes between non-bonded atoms restrict the polypeptide backbone to specific combinations of two dihedral (torsion) angles:

• $\phi$ (Phi): Rotation around the $N - C_\alpha$ bond.
• $\psi$ (Psi): Rotation around the $C_\alpha - C'$ bond.

A Ramachandran Plot graphs $\phi$ versus $\psi$ across all non-glycine residues in a structure. High-quality crystallographic structures exhibit $\ge 98\%$ of residues in the core "favored" regions ($lpha$-helices at $\phi \approx -60^\circ, \psi \approx -45^\circ$; $\beta$-sheets at $\phi \approx -120^\circ, \psi \approx +120^\circ$), with zero non-glycine outliers.

4. Production PyMOL Automation Script: Binding Pocket & Contact Surface Inspection

#!/usr/bin/env python3
"""
Day 13 Practical: PyMOL Automated Structural Analysis Script
Loads PDB structure, highlights binding pocket residues within 4.0 Angstroms,
and renders ray-traced publication figures.
"""

# To run in PyMOL: pymol -c script.py
try:
    from pymol import cmd
except ImportError:
    print("Run inside PyMOL environment (e.g., via PyMOL GUI or python with pymol module).")

def render_binding_site_figure(pdb_id="1tup"):
    """Automate structural rendering of p53-DNA complex."""
    cmd.reinitialize()
    cmd.fetch(pdb_id, type="cif")
    cmd.bg_color("white")
    
    # Protein cartoon representation
    cmd.hide("everything")
    cmd.show("cartoon", "polymer.protein")
    cmd.color("slate", "polymer.protein")
    
    # DNA stick representation
    cmd.show("sticks", "polymer.nucleic")
    cmd.color("orange", "polymer.nucleic and name C*")
    
    # Select critical zinc ion and coordinating cysteines/histidines
    cmd.show("spheres", "name ZN")
    cmd.color("gray70", "name ZN")
    cmd.set("sphere_scale", 0.6, "name ZN")
    
    # Active site contact selection (residues within 3.8 Angstroms of DNA)
    cmd.select("dna_contacts", "polymer.protein within 3.8 of polymer.nucleic")
    cmd.show("sticks", "dna_contacts")
    cmd.color("firebrick", "dna_contacts and name C*")
    
    # High-resolution ray-traced output
    cmd.zoom("dna_contacts", 6)
    cmd.set("ray_shadows", 1)
    cmd.png("p53_dna_binding_pocket.png", width=1600, height=1200, dpi=300, ray=1)
    print("Saved high-resolution render: p53_dna_binding_pocket.png")

if __name__ == "__main__":
    print("PyMOL script ready for command-line or GUI execution.")