An infrared study of the binding and photodissociation of carbon monoxide in cytochrome ba3 from Thermus thermophilus

Einarsdóttir, Ólöf; Killough, P. M.; Fee, James A.; Woodruff, William H.

doi:10.1016/s0021-9258(19)81627-7

Cited by 77 publications

(61 citation statements)

References 18 publications

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“…The events following photolysis of the CO complexes of bacterial and eukaryotic cytochrome oxidase are qualitatively similar in that CO photodissociated from Fe032+ binds to nearby Cub until it is released thermally. The residence time of CO on Cub is longer in the bacterial enzyme, allowing measurement of the absorption spectrm of the CO-Cu vibration using FTIR at ambient temperature (Einarsdóttir et al, 1989). Timeresolved IR measurements on eukaryotic oxidase, in contrast, found a short residence time (half-life of 1.5 µ$) for CO on Cub (Dyer et al, 1989).…”

Section: Resultsmentioning

confidence: 99%

“…The 03(o)-Fe2+ site of the heme-copper oxidases is able to bind a single carbon monoxide molecule that photodissociates from az2+ with high quantum yield and transfers to Cub1+ (Flamingo et al, 1982;Einarsdóttir et al, 1989). The CO complex of eukaryotic oxidases has been used extensively in time-resolved spectral studies as a model for O2 binding and as a photolytic source of unliganded enzyme for reaction with dissolved oxygen.…”

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confidence: 99%

“…However, the results of several recent studies reveal that photodissociation of the CO complex is a more complicated event than previously believed. An IR study of cytochrome baz-CO showed that photodissociation of the Fe-CO bond leads to quantitative binding of CO with Cub (Einarsdóttir et al, 1989). Time-resolved IR studies of cytochrome aaz-CO photolysis at room temperature have directly observed binding of CO to Cub after photolysis (Dyer et al, 1991) and later decay of the Cu-CO complex with a 1.5-ms half-life (Dyer et al, 1989).…”

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confidence: 99%

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Magnetic circular dichroism study of cytochrome ba3 from Thermus thermophilus: spectral contributions from cytochromes b and a3 and nanosecond spectroscopy of carbon monoxide photodissociation intermediates

Goldbeck

Einarsdóttir²,

Dawes³

et al. 1992

Biochemistry

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Near-UV-vis magnetic and natural circular dichroism (MCD and CD) spectra of oxidized, reduced, and carbonmonoxy-complexed cytochrome ba3, a terminal oxidase from the bacterium Thermus thermophilus, and nanosecond time-resolved MCD (TRMCD) and CD (TRCD) spectra of the unligated species formed after photodissociation of the CO complex are presented. The spectral contributions of individual cytochromes b and a3 to the Soret region MCD are identified. TRMCD spectroscopy is used to follow the spin state change (S = 0 to S = 2) in cytochrome a3(2+) following photodissociation of the CO complex. There is prompt appearance of the high-spin state after photolysis, as found previously in mammalian cytochrome oxidase [Goldbeck, R. A., Dawes, T. D., Einarsdóttir, O., Woodruff, W. H., & Kliger, D. S. (1991) Biophys. J. 60, 125-134]. Peak shifts of 1-10 nm appear in the TRMCD, TRCD, and time-resolved UV-vis absorption spectra of the photolyzed enzyme throughout its observable lifetime, indicating that the photolyzed enzyme does not relax to its equilibrium deliganded form before recombination with CO occurs hundreds of milliseconds later. Direct heme-heme interaction is not found in cytochrome ba3, but red-shifts in the MCD and absorption spectra of both cytochromes b and (photolyzed) a3 are correlated with a CO-liganded form of the protein. The long time (tau approximately greater than 1 s) needed for relaxation of the cytochrome b and a3 peaks to their static positions suggests that CO binding to a3 induces a global conformational change in the protein that weakly perturbs the MCD and absorption spectra of b and photolyzed a3. Fea3 binds CO more weakly in cytochrome ba3 than in cytochrome aa3. The MCD spectrum of reduced enzyme solution placed under 1 atm of CO contains a peak at 446 nm that shows approximately 30% of total cytochrome a3 remains pentacoordinate, high-spin.

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Section: Resultsmentioning

confidence: 99%

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confidence: 99%

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confidence: 99%

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Magnetic circular dichroism study of cytochrome ba3 from Thermus thermophilus: spectral contributions from cytochromes b and a3 and nanosecond spectroscopy of carbon monoxide photodissociation intermediates

Goldbeck

Einarsdóttir²,

Dawes³

et al. 1992

Biochemistry

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“…Multiple CO bands arising from different populations of active-site conformers were also observed in flash photolysis studies of CO binding to cytochrome c oxidase. 56,57 Cytochrome c oxidase (CcO) is the terminal respiratory enzyme in the mitochondrial electron transfer chain responsible for reduction of molecular oxygen to water. In strongly visible light, CO is dissociated from the heme a3 pocket and rebinds to Cu B with at least two CO stretching frequencies between 2060 and 2035 cm −1 .…”

Section: ■ Discussionmentioning

confidence: 99%

Substrate-Induced Carbon Monoxide Reactivity Suggests Multiple Enzyme Conformations at the Catalytic Copper M-Center of Peptidylglycine Monooxygenase

Kline

Blackburn

2016

Biochemistry

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The present study uses CO as a surrogate for oxygen to probe how substrate binding triggers oxygen activation in peptidylglycine monooygenase (PHM). Infrared stretching frequencies (ν(C≡O)) of the carbonyl (CO) adducts of copper proteins are sensitive markers of Cu(I) coordination, and are useful in probing oxygen reactivity since the electronic properties of O2 and CO are similar. The carbonyl chemistry has been explored using PHM WT and a number of active site variants in the absence and presence of peptidyl substrates. We have determined that upon carbonylation (i) a major CO band at 2092 cm−1 and a second minor CO band at 2063 cm−1 are observed in the absence of peptide substrate Ac-YVG; (ii) the presence of peptide substrate amplifies the minor CO band and causes it to partially interconvert with the CO band at 2092 cm−1; (iii) the substrate induced CO band is associated with a second conformer at CuM; and (iv) the CuH-site mutants which are inactive, fail to generate any substrate-induced CO bands. The total intensity of both bands is constant suggesting that the Cu(I)M site partitions between the two carbonylated enzyme states. Together, these data provide evidence for two conformers at CuM, one of which is induced by binding of the peptide substrate, with the implication that this represents the conformation that also allows binding and activation of O2.

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“…[97][98][99] In terminal oxidases, CO dissociates from heme a 3 and coordinates to Cu B . The Cu B -CO complex can be trapped at cryogenic temperatures, and 'dark' minus 'illuminated' FTIR difference spectra exhibit a positive m(CO) from the a 3 -CO complex and a negative m(CO) between 2040-2070 cm −1 characteristic of Cu B -CO. [100][101][102][103] In 1998, Saraste and coworkers attempted to use this approach with P.d. cNOR and reported that FTIR difference spectra could only be observed between dark and continuous illumination at 234 K. 104 The asymmetric absorption changes, centered around 1970 cm −1 , were interpreted in terms of a 'dark' signal at 1977 cm −1 , assigned to heme b 3 -CO, and a 'light' signal at 1963 cm −1 , assigned to Fe B -CO.…”

Section: The Diiron Site Can Accommodate Two Co Moleculesmentioning

confidence: 99%

Spectroscopic characterization of heme iron–nitrosyl species and their role in NO reductase mechanisms in diiron proteins

Moënne‐Loccoz

2007

Nat. Prod. Rep.

121

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Nitric oxide (NO) plays an important role in cell signalling and in the mammalian immune response to infection. On its own, NO is a relatively inert radical, and when it is used as a signalling molecule, its concentration remains within the picomolar range. However, at infection sites, the NO concentration can reach the micromolar range, and reactions with other radical species and transition metals lead to a broad toxicity. Under aerobic conditions, microorganisms cope with this nitrosative stress by oxidizing NO to nitrate (NO 3 − ). Microbial hemoglobins play an essential role in this NO-detoxifying process. Under anaerobic conditions, detoxification occurs via a 2-electron reduction of two NO molecules to N 2 O. In many bacteria and archaea, this NOreductase reaction is catalyzed by diiron proteins. Despite the importance of this reaction in providing microorganisms with a resistance to the mammalian immune response, its mechanism remains ill-defined. Because NO is an obligatory intermediate of the denitrification pathway, respiratory NO reductases also provide resistance to toxic concentrations of NO. This family of enzymes is the focus of this review. Respiratory NO reductases are integral membrane protein complexes that contain a norB subunit evolutionarily related to subunit I of cytochrome c oxidase (CcO). NorB anchors one high-spin heme b 3 and one non-heme iron known as Fe B , i.e., analogous to Cu B in CcO. A second group of diiron proteins with NO-reductase activity is comprised of the large family of soluble flavoprotein A found in strict and facultative anaerobic bacteria and archaea. These soluble detoxifying NO reductases contain a non-heme diiron cluster with a Fe-Fe distance of 3.4 Å and are only briefly mentioned here as a promising field of research. This article describes possible mechanisms of NO reduction to N 2 O in denitrifying NO-reductase (NOR) proteins and critically reviews recent experimental results. Relevant theoretical model calculations and spectroscopic studies of the NO-reductase reaction in heme/copper terminal oxidases are also overviewed.

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An infrared study of the binding and photodissociation of carbon monoxide in cytochrome ba3 from Thermus thermophilus

Cited by 77 publications

References 18 publications

Magnetic circular dichroism study of cytochrome ba3 from Thermus thermophilus: spectral contributions from cytochromes b and a3 and nanosecond spectroscopy of carbon monoxide photodissociation intermediates

Magnetic circular dichroism study of cytochrome ba3 from Thermus thermophilus: spectral contributions from cytochromes b and a3 and nanosecond spectroscopy of carbon monoxide photodissociation intermediates

Substrate-Induced Carbon Monoxide Reactivity Suggests Multiple Enzyme Conformations at the Catalytic Copper M-Center of Peptidylglycine Monooxygenase

Spectroscopic characterization of heme iron–nitrosyl species and their role in NO reductase mechanisms in diiron proteins

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