Sulfur donor ligand binding to ferric cytochrome P-450-CAM and myoglobin. Ultraviolet-visible absorption, magnetic circular dichroism, and electron paramagnetic resonance spectroscopic investigation of the complexes.

Sono, Masanori; Andersson, Laura A.; Dawson, John H.

doi:10.1016/s0021-9258(18)34332-1

Cited by 140 publications

(65 citation statements)

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“…In the visible region, the ferric heme exhibits a broad absorption feature with a peak at ∼557 nm and a shoulder at ∼586 nm (Figure A). In previous studies on the cysteine thiolate-coordinated camphor hydroxylase P450cam by Sono et al, the P450 was titrated with the sulfur donor ligand p -chlorothiophenol, generating a ferric bis-Cys heme-ligated model complex, resulting in the formation of a hyperporphyrin spectrum with Soret maxima at 380 and 450 nm . Further studies of the binding of 1-propanethiol to P450cam at pH 6.7 revealed three absorbance peaks in the Soret region at approximately 380, 417, and 465 nm, indicative of a mixture of axial bis-thiolate (the 380 and 465 nm bands) and thiolate/thiol coordination (the 417 nm band).…”

Section: Resultsmentioning

confidence: 97%

“…Further studies of the binding of 1-propanethiol to P450cam at pH 6.7 revealed three absorbance peaks in the Soret region at approximately 380, 417, and 465 nm, indicative of a mixture of axial bis-thiolate (the 380 and 465 nm bands) and thiolate/thiol coordination (the 417 nm band). Increasing the pH resulted in the nearly complete conversion of the 1-propanethiol-bound species to the hyperporphyrin 380 nm/465 nm form at pH 9.1, arising from the deprotonation of the distal ligand to the thiolate state . In studies using porphyrin model complexes, Ullrich et al also reported bis-thiolate complexes with split Soret features and absorption maxima at ∼470 and 380 nm .…”

Section: Resultsmentioning

confidence: 98%

“…In previous studies on the cysteine thiolate-coordinated camphor hydroxylase P450cam by Sono et al, the P450 was titrated with the sulfur donor ligand p-chlorothiophenol, generating a ferric bis-Cys heme-ligated model complex, resulting in the formation of a hyperporphyrin spectrum with Soret maxima at 380 and 450 nm. 43 Further studies of the binding of 1-propanethiol to P450cam at pH 6.7 revealed three absorbance peaks in the Soret region at approximately 380, 417, and 465 nm, indicative of a mixture of axial bis-thiolate (the 380 and 465 nm bands) and thiolate/thiol coordination (the 417 nm band). Increasing the pH resulted in the nearly complete conversion of the 1propanethiol-bound species to the hyperporphyrin 380 nm/465 nm form at pH 9.1, arising from the deprotonation of the distal ligand to the thiolate state.…”

Section: ■ Results and Discussionmentioning

confidence: 99%

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Analysis of Heme Iron Coordination in DGCR8: The Heme-Binding Component of the Microprocessor Complex

et al. 2016

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DGCR8 is the RNA-binding partner of the nuclease Drosha. Their complex (the "Microprocessor") is essential for processing of long, primary microRNAs (pri-miRNAs) in the nucleus. Binding of heme to DGCR8 is essential for pri-miRNA processing. On the basis of the split Soret ultraviolet-visible (UV-vis) spectrum of ferric DGCR8, bis-thiolate sulfur (cysteinate, Cys(-)) heme iron coordination of DGCR8 heme iron was proposed. We have characterized DGCR8 heme ligation using the Δ276 DGCR8 variant and combined electron paramagnetic resonance (EPR), magnetic circular dichroism (MCD), electron nuclear double resonance, resonance Raman, and electronic absorption spectroscopy. These studies indicate DGCR8 bis-Cys heme iron ligation, with conversion from bis-thiolate (Cys(-)/Cys(-)) axial coordination in ferric DGCR8 to bis-thiol (CysH/CysH) coordination in ferrous DGCR8. Pri-miRNA binding does not perturb ferric DGCR8's optical spectrum, consistent with the axial ligand environment being separated from the substrate-binding site. UV-vis absorption spectra of the Fe(II) and Fe(II)-CO forms indicate discrete species exhibiting peaks with absorption coefficients substantially larger than those for ferric DGCR8 and that previously reported for a ferrous form of DGCR8. Electron-nuclear double resonance spectroscopy data exclude histidine or water as axial ligands for ferric DGCR8 and favor bis-thiolate coordination in this form. UV-vis MCD and near-infrared MCD provide data consistent with this conclusion. UV-vis MCD data for ferrous DGCR8 reveal features consistent with bis-thiol heme iron coordination, and resonance Raman data for the ferrous-CO form are consistent with a thiol ligand trans to the CO. These studies support retention of DGCR8 cysteine coordination upon reduction, a conclusion distinct from those of previous studies of a different ferrous DGCR8 isoform.

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Section: Resultsmentioning

confidence: 97%

Section: Resultsmentioning

confidence: 98%

Section: ■ Results and Discussionmentioning

confidence: 99%

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Analysis of Heme Iron Coordination in DGCR8: The Heme-Binding Component of the Microprocessor Complex

et al. 2016

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“…A ferric P450 heme can be high-spin (HS; S = 5 / 2 ) or low-spin (LS; S = 1 / 2 ), and these electronic ground states can be distinguished by optical and EPR spectra. − The UV–vis spectrum of a substrate-bound cytochrome P450 is dependent on the nature of heme ligation. A type I change (blue shift; from ∼418 to ∼390 nm) is associated with an increase in HS character upon ligand binding to the enzyme, whereas a reverse type I spectral change (red shift; from ∼390 to ∼418 nm) reflects a change from more HS to LS character.…”

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confidence: 99%

Biophysical Techniques for Distinguishing Ligand Binding Modes in Cytochrome P450 Monooxygenases

et al. 2020

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The new agreement specifically addresses what authors can do with different versions of their manuscripte.g. use in theses and collections, teaching and training, conference presentations, sharing with colleagues, and posting on websites and repositories. The terms under which these uses can occur are clearly identified to prevent misunderstandings that could jeopardize final publication of a manuscript (Section II, Permitted Uses by Authors).

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“…In some cases, stable models are difficult to generate; for example, assembling thiolate bound heme complexes are synthetically challenging. 68 , 80 − 82 In other cases, reaction of these complexes with the oxidant H 2 O 2 , for example, leads to self-degradation. 83 , 84 Attempting to use O 2 as an oxidant by activating it with ferrous porphyrin requires a chemical reducing agent to reinstate the reduced ferrous heme for catalysis.…”

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confidence: 99%

Rejigging Electron and Proton Transfer to Transition between Dioxygenase, Monooxygenase, Peroxygenase, and Oxygen Reduction Activity: Insights from Bioinspired Constructs of Heme Enzymes

Mukherjee

Dey

2021

JACS Au

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Nature has employed heme proteins to execute a diverse set of vital life processes. Years of research have been devoted to understanding the factors which bias these heme enzymes, with all having a heme cofactor, toward distinct catalytic activity. Among them, axial ligation, distal super structure, and substrate binding pockets are few very vividly recognized ones. Detailed mechanistic investigation of these heme enzymes suggested that several of these enzymes, while functionally divergent, use similar intermediates. Furthermore, the formation and decay of these intermediates depend on proton and electron transfer processes in the enzyme active site. Over the past decade, work in this group, using in situ surface enhanced resonance Raman spectroscopy of synthetic and biosynthetic analogues of heme enzymes, a general idea of how proton and electron transfer rates relate to the lifetime of different O 2 derived intermediates has been developed. These findings suggest that the enzymatic activities of all these heme enzymes can be integrated into one general cycle which can be branched out to different catalytic pathways by regulating the lifetime and population of each of these intermediates. This regulation can further be achieved by tuning the electron and proton transfer steps. By strategically populating one of these intermediates during oxygen reduction, one can navigate through different catalytic processes to a desired direction by altering proton and electron transfer steps.

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Sulfur donor ligand binding to ferric cytochrome P-450-CAM and myoglobin. Ultraviolet-visible absorption, magnetic circular dichroism, and electron paramagnetic resonance spectroscopic investigation of the complexes.

Cited by 140 publications

References 42 publications

Analysis of Heme Iron Coordination in DGCR8: The Heme-Binding Component of the Microprocessor Complex

Analysis of Heme Iron Coordination in DGCR8: The Heme-Binding Component of the Microprocessor Complex

Biophysical Techniques for Distinguishing Ligand Binding Modes in Cytochrome P450 Monooxygenases

Rejigging Electron and Proton Transfer to Transition between Dioxygenase, Monooxygenase, Peroxygenase, and Oxygen Reduction Activity: Insights from Bioinspired Constructs of Heme Enzymes

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