Coordination geometries and vibrational properties of cytochromes a and a3 in cytochrome oxidase from soret excitation Raman spectroscopy

Babcock, Gerald T.; Callahan, Patricia M.; Ondrias, M. R.; Salmeen, Irving T.

doi:10.1021/bi00507a049

Cited by 101 publications

(77 citation statements)

References 47 publications

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“…Upon photolysis of the CO bound species, a competition may occur between O 2 and water binding. Nevertheless these fast rates are consistent with the high spin state found in ferrous deoxy heme a3 in CcO (31,50,51). The present results imply that efficient water pumps must operate in CcO to remove tightly bound molecules of water and to prevent the formation of a low spin state in CcO.…”

Section: Resultssupporting

confidence: 89%

Water may inhibit oxygen binding in hemoprotein models

Collman

Decréau²,

Dey³

et al. 2009

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

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Three distal imidazole pickets in a cytochrome c oxidase (CcO) model form a pocket hosting a cluster of water molecules. The cluster makes the ferrous heme low spin, and consequently the O2 binding slow. The nature of the rigid proximal imidazole tail favors a high spin/low spin cross-over. The O2 binding rate is enhanced either by removing the water, increasing the hydrophobicity of the gas binding pocket, or inserting a metal ion that coordinates to the 3 distal imidazole pickets.spin cross-over ͉ tris-imidazole pocket ͉ water cluster H emoglobin (Hb), myoglobin (Mb) and cytochrome c oxidase (CcO) are hemoproteins that bind O 2, subsequently transporting or reducing it in a 4e-/4Hϩ process (1, 2). The kinetics of oxygen binding to the active sites of these biomolecules are tuned by stabilizing interactions between the oxygen complex and the immediate environment (3, 4). In monometallic proteins such as myoglobin or hemoglobin, a distal histidine stabilizes the oxygen complex by hydrogen bonding (5, 6). A distal copper tris-imidazole ligand in CcO also provides enthalpic stabilization (7). In addition to structural factors affecting oxygen adduct stability in Mb and Hb, it has been suggested that the molecules of water present in the binding pocket could contribute to the relative oxygen affinities (4, 8, 9) in a model dubbed the ''water displacement model.'' This proposed model could logically be extended to CcO where water is the product of the 4e-/4Hϩ reduction of O 2 . Water is expected to be present in larger quantities in CcO than in Hb or Mb, and it is removed from the binding site by water channels (10-13). Here, we describe well-defined biomimetic hemoprotein models 1-4 ( Fig. 1) that demonstrate the role of water in slowing the binding of O 2 . The rates of reaction correlate with the hydrophobicity of the distal pocket (tris-pivalamido-or picket fence in 2, tris-imidazole in 3) and the presence or absence of a distal bound metal [Cu(I) or Zn(II) in 4ab]. Results and DiscussionThe reaction of oxygen with ferrous porphyrins 1-4 was carried out by injecting an anaerobic solution of porphyrin into O 2 -saturated dichloromethane solution (10 mM) under 1 atm of O 2 . Under the initial conditions of the reaction it is assumed, based on earlier studies (15), that the ligand association rate k on (O 2 ) is several orders of magnitude faster than the ligand dissociation rate k off (O 2 ) and consequently the dissociation rate can be assumed negligeable. The O 2 binding rates were obtained by monitoring the change in of the Soret and Q bands in the UV/Vis spectrum (Fig. 2) that shifted from 426 nm to 421 nm, and from 535 nm to 550 nm, respectively. The oxygen complex was characterized at various stages of the reaction by 2 resonance Raman stretches: (i) an oxygen isotope sensitive band observed at 570/544 cm Ϫ1 that corresponds to the Fe-O stretch and (ii) a 4-band (a spin state and redox state marker band) at 1,370 cm Ϫ1 that is typical of a ferric-superoxo species (16)(17)(18)(19). Second order k on (O 2 ) ra...

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Section: Resultssupporting

confidence: 89%

Water may inhibit oxygen binding in hemoprotein models

Collman

Decréau²,

Dey³

et al. 2009

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

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“…Indicated in the figure are the formyl (1610 cm-') and vinyl (1624 cm-') stretches of heme a and the formyl stretch (1662 cm-'), the ring bending mode (365 cm-'), and the Fe-NHis stretch (214 cm-') of heme a3. The spectrum of the wild-type bacterial oxidase is remarkably similar to that of bovine heart cytochrome c oxidase (32)(33)(34)(35), with the notable exception of an increased intensity of the heme a formyl mode at 1610 cm-'. The asterisks indicate the position of the peak due to excess dithionite (deleted).…”

Section: Resultsmentioning

confidence: 79%

Definition of the catalytic site of cytochrome c oxidase: specific ligands of heme a and the heme a3-CuB center.

Shapleigh

Hosler

Tecklenburg

et al. 1992

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

110

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The three-subunit aa3-type cytochrome c oxidase (EC 1.9.3.1) of Rhodobacter sphaeroides is structurally and functionally homologous to the more complex mitochondrial oxidase. The largest subunit, subunit I, is highly conserved and predicted to contain 12 transmembrane segments that provide all the ligands for three of the four metal centers: heme a, heme a3, and CUB. A variety of spectroscopic techniques identify these ligands as histidines. We have used site-directed mutagenesis to change all the conserved histidines within subunit I of cytochrome c oxidase from Rb. sphaeroides. Analysis of the membrane-bound and purified mutant proteins by optical absorption and resonance Raman spectroscopy indicates that His-102 and His-421 are the ligands of heme a, while His-284, His-333, His-334, and His-419 ligate the heme ar-CuB center. To satisfy this ligation assignment, helices II, VI, VII, and X, which contain these histidine residues, must be in close proximity. These data provide empirical evidence regarding the three-dimensional protein structure at the catalytic core of cytochrome c oxidase.Cytochrome c oxidase (EC 1.9.3.1) catalyzes the reduction of oxygen to water at the terminus of the mitochondrial respiratory chain, the principal energy-generating system of eukaryotic organisms (for recent reviews see refs. 1 and 2). Energy conservation is achieved by the coupling of electron transfer through the heme and copper metal centers to proton translocation across the membrane (3). To understand this coupling process it is essential to have a detailed description of the protein structures that form the heme-and copperbinding sites.A number of bacteria synthesize cytochrome c oxidases that are simpler in structure but functionally homologous to the mitochondrial enzymes (2, 4, 5). The aa3-type cytochrome c oxidase from Rhodobacter sphaeroides has recently been purified as a complex of three subunits that are homologues ofthe three mitochondrially encoded subunits of the eukaryotic oxidase (J.P.H. and S.F.-M., unpublished results). The Rb. sphaeroides oxidase has remarkably high turnover (Vm > 1800 sec-1), pumps protons efficiently (0.7 H+/e-) in reconstituted proteoliposomes, and is spectroscopically similar to beef heart cytochrome c oxidase. This preparation clearly establishes the oxidase from Rb. sphaeroides as an excellent bacterial model for examining the structural basis of the catalytic function of the more complex eukaryotic enzyme. The utility of Rb. sphaeroides as a genetic system for analyzing structure/function relationships has been established in previous studies ofthe photosynthetic reaction center (6) and the cytochrome bc complex (7).The genes coding for the three subunits of the Rb. sphaeroides aa3-type oxidase have been cloned and sequenced (refs. 8 and 9 and unpublished data). The genetic organization and deduced amino acid sequences are very similar to those from the closely related bacterium Paracoccus denitrificans (2, 10-12). In addition, subunit I shows 50% sequence identity to subun...

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“…In mammalian cytochrome oxidase and Spirographis-substituted myoglobins, carbonyl groups on the periphery of the hemes result in a pronounced red-shift of the electronic spectra. Carbonyl modes are evident in the spectra of these proteins appearing at 1645-1670 cm-1 [22,23 ] and are quite conformationally sensitive. Since the carbonyl groups on the propionic acids should not be resonantly enhanced, the occurrence of a mode in the 1700 cm -1 region suggests [10] the presence of an a-keto group on a heine substituent or on the chlorin macrocycle.…”

Section: Resultsmentioning

confidence: 99%