Resonance Raman Studies of Co—O2 and O—O Stretching Vibrations in Oxy-Cobalt Hemes

Mackin, Helen C.; Tsubaki, Motonari; Yu, Nai‐Teng

doi:10.1016/s0006-3495(83)84446-4

Cited by 39 publications

(21 citation statements)

References 36 publications

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“…Figure 1 displays the low frequency (50-800 cm -1 ) RR data for 16 O 2 [1A], 18 O 2 [1B], and the normalized difference spectra ( 16 O 2 -18 O 2 ) for wild-type oxyP450cam at 356.4-nm excitation. Numerical results are summarized in Table 1.…”

Section: Resultsmentioning

confidence: 99%

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Identification of the Fe−O−O Bending Mode in Oxycytochrome P450cam by Resonance Raman Spectroscopy

et al. 1998

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An oxygen-sensitive mode in oxygenated wild-type cytochrome P450cam (oxyP450cam) is observed at 401 cm-1 and assigned to the δ(Fe−O−O) bending mode, based upon 16O2,18O2 isotopic shifts (19 cm-1) and comparison with Co− and Fe−oxyporphyrin complexes. The detection of this Fe−O−O bending mode has structural implications for enzyme function since its frequency reflects the energies associated with Fe−O−O distortion in oxyhemeprotein active sites. Three body normal coordinate calculations adequately fit the experimental data set with a 125−130° bond angle for the Fe−O−O linkage in oxyP450cam. Observation of low-frequency isotope-sensitive vibrational patterns, some of which are hypothesized to be associated with out-of-plane porphyrin motions are also reported. These patterns, in conjunction with the high frequency of this bending mode and the abnormal isotopic shift of the Fe−O2 stretching mode, suggest a “strained” Fe−O−O moiety in oxyP450cam, with comparable mobility to HbO2 and MbO2. Possible sources of this “strain” and implications for catalytic dioxygen activation in P450cam are discussed.

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Section: Resultsmentioning

confidence: 99%

“…Oxygenation was performed by the addition of 5 mL of the desired isotopic gas to the sealed 60 µL aliquots which were then transferred to the sample cell, whereupon data collection commenced. 18 O2 was purchased from ICON (Mt. Marion, NY).…”

Section: Methodsmentioning

confidence: 99%

Identification of the Fe−O−O Bending Mode in Oxycytochrome P450cam by Resonance Raman Spectroscopy

et al. 1998

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“…It is interesting to note that these vibrations are pH-insensitive in oxy cobalt myoglobin, which does not exhibit a Bohr effect at room temperature (47). The vibrational modes of the cobalt-dioxygen moiety in the low-pH form of cobalt CTT III are similar to those found for the oxy cobalt picket fence porphyrin (48). These similarities in the ligand-associated vibrations between myoglobin and the high-pH form of CTT III and between picket fence porphyrin and the low-pH form of CTT III, respectively, indicate similar binding geometries for the bound dioxygen in each of these pairs.…”

Section: Discussionmentioning

confidence: 57%

Resonance Raman evidence for the mechanism of the allosteric control of O2-binding in a cobalt-substituted monomeric insect hemoglobin

Thompson¹,

Yu²,

Gersonde³

1987

Biophysical Journal

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The substitution of iron for cobalt in the monomeric insect hemoglobin CTT (Chironomus thummi thummi) III does not alter the Bohr effect for O2-binding. The cobalt substitution in this hemoglobin allows us to identify not only the O-O and Co-O2 stretching mode but also the Co-O-O bending mode by resonance Raman spectroscopy. The assignments were made via 16O2/18O2 isotope exchange. The modes associated with the Co-O-O moiety are pH-dependent. These pH-induced changes of the resonance Raman spectra are correlated with the t = r conformation transition. At high pH (high-affinity state) two unperturbed O-O stretching modes are observed at 1,068 cm-1 (major component) and 1,093 cm-1 (minor component) for the 18O2 complex. These frequencies correspond to split modes at 1,107 cm-1 and 1,136 cm-1 and an unperturbed mode at approximately 1,153 cm-1 for the 16O2 complex. At low pH (low-affinity state) the minor component becomes the major component and vice versa. The Co-O2 stretching frequency varies for approximately 520 cm-1 (pH 5.5) to 537 cm-1 (pH 9.5) indicating a stronger (hence shorter) Co-O2 bond in the high-affinity state. On the other hand, the O-O bond is weakened upon the conversion of the low- to the high-affinity state. The Co-O-O bending mode changes from 390 cm-1 (pH 9.5) to 374 cm-1 (pH 5.5). In the deoxy form the resonance Raman spectra are essentially pH-insensitive except for a vinyl mode at 414 cm-1 (pH 5.5), which is shifted to 416 cm-1 (pH 5.5).

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“…For example, electron paramagnetic resonance (EPR) studies on cobalt-substituted sperm whale myoglobin (CoMb) enabled detailed characterization of the paramagnetic CoMbO 2 complex (3,4,24,25). In analogous oxygenbinding studies in CoMb and cobalt-substituted hemoglobin (5,6,26), resonance Raman was used to identify the O-O stretching mode because cobalt-substituted proteins exhibit enhancement of this vibrational mode compared to the native iron proteins.…”

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confidence: 99%

De novo biosynthesis of a nonnatural cobalt porphyrin cofactor inE. coliand incorporation into hemoproteins

Perkins

Weaver

Buller

et al. 2021

Proc. Natl. Acad. Sci. U.S.A.

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Enzymes that bear a nonnative or artificially introduced metal center can engender novel reactivity and enable new spectroscopic and structural studies. In the case of metal-organic cofactors, such as metalloporphyrins, no general methods exist to build and incorporate new-to-nature cofactor analogs in vivo. We report here that a common laboratory strain, Escherichia coli BL21(DE3), biosynthesizes cobalt protoporphyrin IX (CoPPIX) under iron-limited, cobalt-rich growth conditions. In supplemented minimal media containing CoCl2, the metabolically produced CoPPIX is directly incorporated into multiple hemoproteins in place of native heme b (FePPIX). Five cobalt-substituted proteins were successfully expressed with this new-to-nature cobalt porphyrin cofactor: myoglobin H64V V68A, dye decolorizing peroxidase, aldoxime dehydratase, cytochrome P450 119, and catalase. We show conclusively that these proteins incorporate CoPPIX, with the CoPPIX making up at least 95% of the total porphyrin content. In cases in which the native metal ligand is a sulfur or nitrogen, spectroscopic parameters are consistent with retention of native metal ligands. This method is an improvement on previous approaches with respect to both yield and ease-of-implementation. Significantly, this method overcomes a long-standing challenge to incorporate nonnatural cofactors through de novo biosynthesis. By utilizing a ubiquitous laboratory strain, this process will facilitate spectroscopic studies and the development of enzymes for CoPPIX-mediated biocatalysis.

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Resonance Raman Studies of Co—O2 and O—O Stretching Vibrations in Oxy-Cobalt Hemes

Cited by 39 publications

References 36 publications

Identification of the Fe−O−O Bending Mode in Oxycytochrome P450cam by Resonance Raman Spectroscopy

Identification of the Fe−O−O Bending Mode in Oxycytochrome P450cam by Resonance Raman Spectroscopy

Resonance Raman evidence for the mechanism of the allosteric control of O2-binding in a cobalt-substituted monomeric insect hemoglobin

De novo biosynthesis of a nonnatural cobalt porphyrin cofactor inE. coliand incorporation into hemoproteins

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