The mutation Met121His creates a type-1.5 copper site in Alcaligenes denitrificans azurin

Kroes, S.J.; Hoitink, C.W.G.; Andrew, C.R.; Ai, J.; Sanders-Loehr, J.; Messerschmidt, A.; ... ; Canters, G.W. (1996)

Article / Letter to editor

The Cu ligand Met121 in azurin of Alcaligenes denitrificans was mutated to His. The spectroscopic and mechanistic properties of [M121H]azurin appear to be pH dependent with a pK(a) of 3.8 due to the ionization of His121. The [M121H]azurin mutant exhibits two major distinct metal-site-coordination geometries which coexist in solution according to a pH-dependent equilibrium. Both species have been spectroscopically characterized by ultraviolet-visible, EPR and resonance Raman spectroscopies. At neutral DH. His121 is deprotonated and acts as the fourth ligand of the Cu; the spectroscopic characteristics of the Cu site at this pH are halfway between those of a type-1 and a type-2 Cu site, and the site is referred to as a type-1.5 or intermediate Cu site. The spectral data are compatible with a tetrahedral geometry of this site. At low pH, the spectroscopic data indicate that [M121H]azurin has a trigonal type-1 rhombic Cu site.

• All authors: Kroes, S.J.; Hoitink, C.W.G.; Andrew, C.R.; Ai, J.; Sanders-Loehr, J.; Messerschmidt, A.; Hagen, W.R.; Canters, G.W.
• Date: 1996-09-01
• Journal: European Journal of Biochemistry
• Volume: 240
• Issue: 2
• Pages: 342 - 351
• DOI: doi:10.1111/j.1432-1033.1996.0342h.x