Engineering His(E7) affects the control of heme reactivity in Aplysia limacina myoglobin

Aplysia limacina myoglobin lacks the distal histidine (His (E7)) and displa ys a ligand stabilization mechanism based on Arg(E10). The double mutant Va l(E7)His-Arg(E10)Thr has been prepared to engineer the role of His(E7), typ ical of mammalian myoglobins, in a different globin framework. The 2.0 Angs trom crystal structure of Val(E7)His-Arg(E10)Thr met-Mb mutant reveals that the His(E7) side chain points out of the distal pocket, providing an expla nation for the observed failure to stabilize the Fe(II) bound oxygen in the ferrous myoglobin. Moreover, spectroscopic analysis together with kinetic data on azide binding to met-myoglobin are reported and discussed in terms of the presence of a water molecule at coordination distance from the heme iron, (C) 2000 Academic Press.