Blocking the gate to ligand entry in human hemoglobin.

摘要: His(E7) to Trp replacements in HbA lead markedly biphasic bimolecular CO rebinding after laser photolysis. For isolated mutant subunits, the fraction of fast phase increases with increasing [CO], suggesting a competition between binding an open conformation empty E7 channel and relaxation blocked or closed, slowly reacting states. The rate conformational state is ∼18,000 s−1 α subunits ∼10-fold faster β ∼175,000 s−1. Crystal structures were determined for tetrameric α(WT)β(Trp-63) HbCO, α(Trp-58)β(WT) deoxyHb, Trp-64 deoxy- CO-Mb as controls. In Trp-63(E7) βCO, indole side chain located solvent interface, blocking entry into channel. Similar Trp-64(E7) conformations are observed Mb crystal structures. Trp-58(E7) deoxy-α fills both distal pocket, forming completely closed state. constant binding, k′CO, Hb ∼80–90 μm−1 s−1, whereas k′CO states 1000-fold slower, ∼0.08 A transient intermediate ≈ 0.7 photolysis βCO indicates that ring blocks entrance channel, Trp(E7) deoxyMb subunits. Thus, either filling dramatically slows providing strong evidence major pathway (≥90%) ligand human hemoglobin.