PDBsum entry 1aeq

Oxidoreductase

PDB id: 1aeq

Contents

Protein chain

291 a.a.

Ligands

HEM

2EZ

Waters ×100

References listed in PDB file

Key reference

• Title: Artificial protein cavities as specific ligand-Binding templates: characterization of an engineered heterocyclic cation-Binding site that preserves the evolved specificity of the parent protein.
• Authors: R.A.Musah, G.M.Jensen, S.W.Bunte, R.J.Rosenfeld, D.B.Goodin.
• Ref.: J Mol Biol, 2002, 315, 845-857.
• PubMed id: 11812152

Abstract

Cavity complementation has been observed in many proteins, where an appropriate small molecule binds to a cavity-forming mutant. Here, the binding of compounds to the W191G cavity mutant of cytochrome c peroxidase is characterized by X-ray crystallography and binding thermodynamics. Unlike cavities created by removal of hydrophobic side-chains, the W191G cavity does not bind neutral or hydrophobic compounds, but displays a strong specificity for heterocyclic cations, consistent with the role of the protein to stabilize a tryptophan radical at this site. Ligand dissociation constants for the protonated cationic state ranged from 6 microM for 2-amino-5-methylthiazole to 1 mM for neutral ligands, and binding was associated with a large enthalpy-entropy compensation. X-ray structures show that each of 18 compounds with binding behavior bind specifically within the artificial cavity and not elsewhere in the protein. The compounds make multiple hydrogen bonds to the cavity walls using a subset of the interactions seen between the protein and solvent in the absence of ligand. For all ligands, every atom that is capable of making a hydrogen bond does so with either protein or solvent. The most often seen interaction is to Asp235, and most compounds bind with a specific orientation that is defined by their ability to interact with this residue. Four of the ligands do not have conventional hydrogen bonding atoms, but were nevertheless observed to orient their most polar CH bond towards Asp235. Two of the larger ligands induce disorder in a surface loop between Pro190 and Asn195 that has been identified as a mobile gate to cavity access. Despite the predominance of hydrogen bonding and electrostatic interactions, the small variation in observed binding free energies were not correlated readily with the strength, type or number of hydrogen bonds or with calculated electrostatic energies alone. Thus, as with naturally occurring binding sites, affinities to W191G are likely to be due to a subtle balance of polar, non-polar, and solvation terms. These studies demonstrate how cavity complementation and judicious choice of site can be used to produce a protein template with an unusual ligand-binding specificity.

Secondary reference #1

• Title: Small molecule binding to an artificially created cavity at the active site of cytochrome c peroxidase.
• Authors: M.M.Fitzgerald, M.J.Churchill, D.E.Mcree, D.B.Goodin.
• Ref.: Biochemistry, 1994, 33, 3807-3818. [DOI no: 10.1021/bi00179a004]
• PubMed id: 8142383

Secondary reference #2

• Title: The asp-His-Fe triad of cytochrome c peroxidase controls the reduction potential, Electronic structure, And coupling of the tryptophan free radical to the heme.
• Authors: D.B.Goodin, D.E.Mcree.
• Ref.: Biochemistry, 1993, 32, 3313-3324. [DOI no: 10.1021/bi00064a014]
• PubMed id: 8384877