Normal coordinate analysis of the copper center of azurin and the assignment of its resonance Raman spectrum.

Abstract

Normal coordinate analysis that utilizes a general valence force field and the Wilson FG matrix method has been applied to several structural models representing the active site of the blue copper protein, azurin. The models included tetrahedral and square planar CuN2SS', trigonal CuN2S, and trigonal bipyramidal CuN2SS'O structures in which the Ns are imidazole nitrogens of histidines, S is the thiolate sulfur ofcysteine, S' is the thioether sulfur ofmethionine, and 0 is a peptide carbonyl oxygen. For constant Cu-ligand bond lengths and initial force constants, the force field was refined against the most intense ofthe observed frequencies (424, 404, 369, and 261 cm-) in the resonance Raman spectrum of Pseudomonas aeruginosa azurin. The most satisfactory fit between observed and calculated frequencies occurs for tetrahedral and trigonal structures. The calculations provide detailed assignments for the resonance Raman spectrum of azurin and reveal considerable mixing of Cu-S(Cys) and Cu-N(His) vibrational modes. The trigonal model is favored because it is shown that the =260-cm-vibration is an invariant feature in the resonance Raman spectra ofblue copper proteins, even those lacking a methionine in the vicinity of the copper atom. The present analysis ascribes the high frequencies ofthe Cu-ligand stretching modes and the resonance enhancement to the coupled nature of their vibrations and the Franck-Condon overlaps with predominant (Cys)S--Cu(II) charge transfer bands in the visible region.

Temperature dependence of the resonance Raman spectra of plastocyanin and azurin between cryogenic and ambient conditions.

“…with a medium-intensity peak near 265 cm-' and weaker, poorly resolved features elsewhere (11)(12)(13)(14)(15)23). Franck-Condon arguments have led investigators in the past to conclude that the intense RR peaks are due to the stretching vibrations of the stronger copper-ligand bonds (11)(12)(13)(14)23).…”

“…Franck-Condon arguments have led investigators in the past to conclude that the intense RR peaks are due to the stretching vibrations of the stronger copper-ligand bonds (11)(12)(13)(14)23). It has commonly been assumed that such motions are likely to be enhanced by strong coupling to the resonant electronic chromophore-i.e., the ligand-to-metal charge-transfer transitions involving copper(II)-bound cysteine sulfur.…”

“…It has commonly been assumed that such motions are likely to be enhanced by strong coupling to the resonant electronic chromophore-i.e., the ligand-to-metal charge-transfer transitions involving copper(II)-bound cysteine sulfur. The peak near 265 cm-' has generally been taken to be a stretching vibration involving a more weakly bound ligand (11)(12)(13)(14) although other assignments have been suggested (23).…”

Conformational substates in azurin.

“…Extensive studies of the physical and chemical properties of the copper site have been carried out (12). The copper center that gives rise to the intense blue color ofthe protein has been characterized by optical (13), electron paramagnetic resonance (EPR) (14)(15)(16), NMR (17), and Raman spectroscopy (18). Blue copper centers (type 1 copper) bind nitric oxide (NO) (19).…”

Ultrafast excited-state charge-transfer dynamics in laccase type I copper site

“…In particular, intense peaks , which constitute in general the fingerprint of the RR spectra of T1 copper sites, can be assigned to the mixing of the Cu-S(Cys) stretching vibration with multiple heavy atom bending modes of the ligand and adjacent residues [30][31][32][33]. POXC RR vibrational features are rather similar to those observed in some laccases and blue copper proteins, characterized by similar organization of the copper active site [30][31][32][33][34]. Slight modulations in the positions of the main peaks could be related to differences in the T1 copper site as well as in the surrounding protein matrix.…”