Resonance Raman Investigation of Imidazole and Imidazolate Complexes of Microperoxidase: Characterization of the Bis(histidine) Axial Ligation in c-Type Cytochromes

Abstract

In order to evaluate the steric and electronic influences of the heme axial ligands on the vibrational modes of heme c, various ferric and ferrous six-coordinate heme c compounds have been prepared from microperoxidase-8 (MP8) and different extrinsic ligands. In this paper, the absorption and Soret-excited resonance Raman (RR) spectra of imidazole, imidazolate, 1-methylimidazole, and histidine complexes of MP8 are presented. The absorption characteristics of the unligated forms, either aggregated or monomeric, as well as of the ligated forms of MP8(III) and MP8(II) have been determined as a function of pH, the presence of a cationic detergent, and the ligand concentration. Spectrophotometric titrations have shown that MP8(III) and MP8(II) can bind one or two molecules of exogenous ligand, forming monoligated or bisligated complexes. The latter form, observed with large excesses of ligand, results from the displacement of the intrinsic proximal His of MP8 by an exogenous ligand. Several structural marker bands have been detected in the high- and low-frequency regions of RR spectra. The high-frequency RR spectra of the ImH compounds of MP8(III) exhibit a v10 mode sensitive to ligand deprotonation(s). Moreover, the replacement of His by an exogenous ImH in MP8(III) complexes induces the upshift of the v10 mode frequency (1637-1641 cm-1), indicating that the porphyrin skeleton is less distorted when the internal coordination of proximal His to heme is broken. A similar dependence of the out-of-plane porphyrin distortion is suggested for the low-frequency mode v8 (343-347 cm-1). As far as the ferrous compounds are concerned, the mode most sensitive to the ImH deprotonation is v11, which is downshifted from 1539 to 1527 cm-1. Comparisons of the low-frequency regions of the RR spectra of imidazole-type ligated MP8(III) and MP8(II) complexes, as well as observations based on isotopic substitutions of the corresponding 1-methylimidazole complexes (MeIm-->MeIm-d6), allow the assignment of two bands in the 184-197 and 400-409 cm-1 regions to modes involving the symmetric and asymmetric stretches of the axial ligands, respectively. Two other bands in the 343-347 and 359-362 cm-1 regions, sensitive to the mass and/or deprotonation states of the axial ligands, have been tentatively assigned to v(Fe-N(pyrrole)) modes coupled to either a deformation mode of axial bonds or an internal mode of the bound imidazole(s).(ABSTRACT TRUNCATED AT 400 WORDS)

Structural and EPR Characterization of the Soluble Form of Cytochrome c-550 and of the psbV2 Gene Product from the Cyanobacterium Thermosynechococcus elongatus

“…In particular, the n 8 and n 50 modes are observed at 347 and 364 cm -1 , respectively. Considering previous RR data on various ferric cytochromes and heme model compounds, the high-and lowfrequency regions of RR spectra of the psbV2 gene product are characteristic of a six-coordinated low-spin c-type heme (Desbois 1994, Döpner et al 1998, Hu et al 1993, Kitagawa et al 1975, Lefevre-Groboillot et al 2001, Othman and Desbois 1998, Othman et al 1994, Othman et al 1997, Verma et al 1988. However, the observed frequencies for the n 2 , n 3 , n 4 , n 8 , n 10 and n 50 modes are systematically higher when compared to those previously published for neutral ferric cyt c, cyt c 2 (His/ Met coordination) or cyt c 3 (His/His coordination) (Hu et al 1993, Kitagawa et al 1975, Othman et al 1997, Verma et al 1988.…”

“…Among the available vibrational data, only the RR frequencies of an alkaline form of cyt c (His/hydroxide coordination) closely resemble those of the psbV2 gene product (Döpner et al 1998). Ferric microperoxidase-8 [MP8(III)] was previously used to model different axial coordination of heme c (Othman and Desbois 1998, Othman et al 1994, LefevreGroboillot et al 2001. RR studies on six-coordinated low-spin MP8(III) complexes showed that n 4 , n 3 , n 2 , n 10 , n 8 and n 50 are sensitive to the axial heme ligation (Othman and Desbois 1998) (Table 3).…”

The Critical Role of the Proximal Calcium Ion in the Structural Properties of Horseradish Peroxidase

“…The same experiment was also carried out with 413.1 nm excitation, where the LS form is selectively resonance enhanced (data not shown). For both excitation wavelengths (441.6 and 413.1 nm) the spectrum of Ca 2ϩ -depleted HRPC at pH 7.8 (identical to that at pH 9.6) appears similar to that of the HRP⅐Im complex and, hence, contains a significant proportion of a bis-histidyl 6-c LS heme (25,37,38). Nevertheless, in accord with the absorption spectra, the presence of two 3 bands at 1471 and 1492 cm Ϫ1 , corresponding to HS and LS species, respectively, together with the relative intensity of the bands at 1556 and 1580 cm Ϫ1 (corresponding essentially to 2 bands of HS and LS species, respectively) indicate that in Ca 2ϩ -depleted HRPC an additional minor HS component is also present.…”

Factors that determine the unusually low reduction potential of cytochrome c 550 in cyanobacterial photosystem II

“…It is typical of c-type cytochromes and consistent with a six-coordinate low-spin (6c,ls) ferric heme with bis-His axial ligation. A comparison of the vibrational modes of cyt c 550 with those of the bis(N-methylimidazole) complex of ferric protoporphyrin IX dimethyl ester [Fe(III)PPIX-DME] [22] and the imidazole adduct of ferric microperoxidase-8 [MP8(III)] [23] is presented in Table 1. The vibrational modes are assigned on the basis of the normal coordinate analysis of isotopically labeled cyt c [24].…”