title: Determination of ligand pathways in globins: apolar tunnels versus polar gates.
abstract: Although molecular dynamics simulations suggest multiple interior pathways for O(2) entry into and exit from globins, most experiments indicate well defined single pathways. In 2001, we highlighted the effects of large-to-small amino acid replacements on rates for ligand entry and exit onto the three-dimensional structure of sperm whale myoglobin. The resultant map argued strongly for ligand movement through a short channel from the heme iron to solvent that is gated by the distal histidine (His-64(E7)) near the solvent edge of the porphyrin ring. In this work, we have applied the same mutagenesis mapping strategy to the neuronal mini-hemoglobin from Cerebratulus lacteus (CerHb), which has a large internal tunnel from the heme iron to the C-terminal ends of the E and H helices, a direction that is 180° opposite to the E7 channel. Detailed comparisons of the new CerHb map with expanded results for Mb show unambiguously that the dominant (>90%) ligand pathway in CerHb is through the internal tunnel, and the major (>75%) ligand pathway in Mb is through the E7 gate. These results demonstrate that: 1) mutagenesis mapping can identify internal pathways when they exist; 2) molecular dynamics simulations need to be refined to address discrepancies with experimental observations; and 3) alternative pathways have evolved in globins to meet specific physiological demands.
authors.affiliation: Department of Biochemistry and Cell Biology, Rice University, Houston, Texas 77005-1892, USA.
authors.forename: Mallory D
authors.initials: MD
authors.lastname: Salter
authors.forename: George C
authors.initials: GC
authors.lastname: Blouin
authors.forename: Jayashree
authors.initials: J
authors.lastname: Soman
authors.forename: Eileen W
authors.initials: EW
authors.lastname: Singleton
authors.forename: Sylvia
authors.initials: S
authors.lastname: Dewilde
authors.forename: Luc
authors.initials: L
authors.lastname: Moens
authors.forename: Alessandra
authors.initials: A
authors.lastname: Pesce
authors.forename: Marco
authors.initials: M
authors.lastname: Nardini
authors.forename: Martino
authors.initials: M
authors.lastname: Bolognesi
authors.forename: John S
authors.initials: JS
authors.lastname: Olson
mesh_terms: D000818:Animals; D003062:Codon; D018360:Crystallography, X-Ray / Q000379:methods; D006454:Hemoglobins / Q000737:chemistry*; D007448:Invertebrates / Q000235:genetics / Q000378:metabolism*; D008024:Ligands; D008958:Models, Molecular; D008968:Molecular Conformation; D056004:Molecular Dynamics Simulation; D009154:Mutation; D009211:Myoglobin / Q000737:chemistry*; D009569:Nitric Oxide / Q000737:chemistry; D010100:Oxygen / Q000737:chemistry*; D011994:Recombinant Proteins / Q000737:chemistry; D012997:Solvents / Q000737:chemistry; D050661:Sperm Whale
chemical_list: D003062:Codon; D006454:Hemoglobins; D008024:Ligands; D009211:Myoglobin; D011994:Recombinant Proteins; D012997:Solvents; D009569:Nitric Oxide; D010100:Oxygen