title: Spectroscopic and electrochemical properties of two azurins (Az-iso1 and Az-iso2) from the obligate methylotroph Methylomonas sp. strain J and the structure of novel Az-iso2.
abstract: Two azurin-type blue copper proteins, which are related to the electron-transfer processes involving methylamine/methanol oxidation, have been spectroscopically and electrochemically characterized. The obligate methylotroph Methylomonas sp. strain J gives rise to two azurins (Az-isol and Az-iso2) with methylamine dehydrogenase (MADH-Mj). The intense blue bands characteristic of Az-iso1 and Az-iso2 are observed at 621 and 616 nm in the visible absorption spectra respectively, being revealed at 620-630 nm in those of usual azurins. The EPR signal of Az-iso1, similar to usual azurins, shows axial symmetry, while the axial EPR signal of Az-iso2 involves a slightly rhombic character. The half-wave potentials (E1/2) of the two azurins and the intermolecular electron-transfer rate constants (kET) from MADH-Mj to each azurin were determined by cyclic voltammetry. The E1/2 values of Az-iso1 and Az-iso2 are +321 and +278 mV vs NHE at pH 7.0, respectively. The kET value of Az-iso2 is larger than that of Az-iso1 by a factor of 5. However, the electron-transfer rate of Az-iso2 is interestingly slower than those of the azurins from a denitrifying bacterium, Alcaligenes xylosoxidans NCIB 11015, and the amicyanin from a different methylotroph, Methylobacterium extorquens AM1. The structure of Az-iso2 has been determined and refined against 1.6 A X-ray diffraction data. The whole structure of Az-iso2 is quite similar to those of azurins reported already. The Cu(II) site of Az-iso2 is a distorted trigonal bipyramidal geometry like those of other azurins, but some of the Cu-ligand distances and ligand-Cu-ligand bond angle parameters are slightly different. These findings suggest that Az-iso2 is a novel azurin and perhaps functions as an electron acceptor for MADH.
authors.affiliation: Department of Chemistry, Graduate School of Science, Osaka University, Japan. bic@ch.wani.osaka-u.ac.jp
authors.forename: S
authors.initials: S
authors.lastname: Suzuki
authors.forename: N
authors.initials: N
authors.lastname: Nakamura
authors.forename: K
authors.initials: K
authors.lastname: Yamaguchi
authors.forename: K
authors.initials: K
authors.lastname: Kataoka
authors.forename: T
authors.initials: T
authors.lastname: Inoue
authors.forename: N
authors.initials: N
authors.lastname: Nishio
authors.forename: Y
authors.initials: Y
authors.lastname: Kai
authors.forename: J
authors.initials: J
authors.lastname: Tobari
mesh_terms: D001400:Azurin / Q000737:chemistry*; D002942:Circular Dichroism; D004563:Electrochemistry; D004578:Electron Spin Resonance Spectroscopy; D020620:Methylomonas / Q000737:chemistry*; D008958:Models, Molecular; D011487:Protein Conformation; D020033:Protein Isoforms / Q000737:chemistry*
chemical_list: D020033:Protein Isoforms; D001400:Azurin