Cobalt-cytochrome c. I. Preparation, properties, and enzymic activity

Dickinson, L. Charles; Chien, James C. W.

doi:10.1021/bi00687a003

Cited by 64 publications

(46 citation statements)

References 39 publications

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“…This protein species was stable for several hours as judged from the visible spectrum. The spectrum was indicative of a low spin state of the cobaltic ion, which is consistent with the chemical property of this metal (25,26). No modification of the Co-PPIX was observed as judged from the spectra including the pyridine addition (metallochrome) spectra (Table I).…”

Section: Table I Absorption Maxima Of Various Species Derived From Ofsupporting

confidence: 67%

In Vitro Studies on Thioether Bond Formation between Hydrogenobacter thermophilus Apocytochrome c552 with Metalloprotoporphyrin Derivatives

Daltrop

Ferguson

2004

Journal of Biological Chemistry

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Section: Table I Absorption Maxima Of Various Species Derived From Ofsupporting

confidence: 67%

In Vitro Studies on Thioether Bond Formation between Hydrogenobacter thermophilus Apocytochrome c552 with Metalloprotoporphyrin Derivatives

Daltrop

Ferguson

2004

Journal of Biological Chemistry

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“…Cobalt-Substituted Cyt C. The absorption spectra of horse, tuna, and yeast Co-Cyt c are virtually identical (Soret maximum, 427 nm; Q-bands, 534, 568 nm), indicating that their heme environments are the same (30,31). The far-UV CD spectrum of Co-Cyt c displays minima at 208 and 222 nm, characteristic of ␣-helical structures in the folded protein.…”

Section: Resultsmentioning

confidence: 99%

“…Fluorescence decay kinetics were measured as described previously (27). Horse, tuna, and yeast Co-Cyt were prepared according to established protocols (30,31) with minor modifications. For the modification of yeast-Co-Cyt c with a dansyl fluorophore at Cys-102 (DNS-Co-Cyt c), FPLC-purified yeast Co-Cyt c was treated with excess 5-({[2-iodoacetyl)amino]ethyl}amino)-naphthalene-1-sulfonic acid (IAEDANS) (Molecular Probes), a thiol-specific derivative of the dansyl-fluorophore.…”

Section: Methodsmentioning

confidence: 99%

Using deeply trapped intermediates to map the cytochrome c folding landscape

Tezcan

Findley

Crane

et al. 2002

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

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Replacement of iron with cobalt(III) selectively introduces a deep trap in the folding-energy landscape of the heme protein cytochrome c. Remarkably, neither the protein structure nor the folding thermodynamics is perturbed by this metal-ion substitution, as shown by data from spectroscopic and x-ray diffraction experiments. Through kinetics measurements, we have found parallel folding pathways involving several different misligated Co(III) species, and, as these folding intermediates persist for several hours under certain conditions, we have been able to elucidate fully their spectroscopic properties. The results, along with an analysis of the fluorescence energy-transfer kinetics during refolding, show that rapidly equilibrating populations of compact and extended polypeptide conformations are present until all molecules have reached the native structure. These measurements provide direct evidence that collapsed denatured structures are not substantially more stable than extended conformations of cytochrome c. Significant advances in theory and experiment are producing an increasingly detailed description of how a polypeptide self-assembles into the native structure of a protein (1-11). Energy landscape theories (1, 12-15), in particular, have provided a framework for interpreting experimental investigations of protein folding kinetics. Despite this progress, many fundamental questions about protein folding dynamics remain to be answered.Years of experimental work on the energetics and dynamics of self-assembly of cytochrome c (Cyt c) have failed to resolve whether unfolded polypeptides undergo global collapse to compact conformations on dilution or laser triggering to solution conditions that strongly favor the native structure (11,(16)(17)(18)(19)(20)(21)(22)(23)(24)(25)(26). Indeed, recent kinetics experiments suggest that comparable populations of compact and extended polypeptides are formed rapidly (Ͻ1 ms), and these two populations disappear in parallel as the protein folds (27,28). To test this energy landscape model rigorously, we have isolated the structures that comprise the submillisecond (or ''burst'') phase by replacing the native iron center with cobalt(III) (29): because of very high ligand substitution barriers, the final step of cobalt(III)-Cyt c (Co-Cyt c) folding is well separated in time from all early events, and, in this kinetically stabilized system, the early folding intermediates can be examined carefully over a period of hours by an entire armamentarium of physical methods. Materials and MethodsUV-visible and CD and fluorescence spectra were collected with Hewlett-Packard 8452A, Aviv (Lakewood, NJ) 62A DS, and Hitachi (Tokyo) F-4500 spectrometers, respectively. Fluorescence decay kinetics were measured as described previously (27). Horse, tuna, and yeast Co-Cyt were prepared according to established protocols (30, 31) with minor modifications. For the modification of yeast-Co-Cyt c with a dansyl fluorophore at Cys-102 (DNS-Co-Cyt c), FPLC-purified yeast Co-Cyt c was treated with exce...

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“…While replacement of the central Fe in horse heart cytochrome c with Zn (54), Co (16), and Sn (54) by in vitro manipulation has been demonstrated, the use of alternative metal porphyrins by cytochrome c biogenesis systems has not been tested. Here various methods to detect ZnPPIX (and SnPPIX) insertion into apocytochrome c by either the system I or II cytochrome c biogenesis pathway were employed.…”

Section: Discussionmentioning

confidence: 99%

Heme Concentration Dependence and Metalloporphyrin Inhibition of the System I and II CytochromecAssembly Pathways

et al. 2007

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Studies have indicated that specific heme delivery to apocytochrome c is a critical feature of the cytochrome c biogenesis pathways called system I and II. To determine directly the heme requirements of each system, including whether other metal porphyrins can be incorporated into cytochromes c, we engineered Escherichia coli so that the natural system I (ccmABCDEFGH) was deleted and exogenous porphyrins were the sole source of porphyrins (⌬hemA). The engineered E. coli strains that produced recombinant system I (from E. coli) or system II (from Helicobacter) facilitated studies of the heme concentration dependence of each system. Using this exogenous porphyrin approach, it was shown that in system I the levels of heme used are at least fivefold lower than the levels used in system II, providing an important advantage for system I. Neither system could assemble holocytochromes c with other metal porphyrins, suggesting that the attachment mechanism is specific for Fe protoporphyrin. Surprisingly, Zn and Sn protoporphyrins are potent inhibitors of the pathways, and exogenous heme competes with this inhibition. We propose that the targets are the heme binding proteins in the pathways (CcmC, CcmE, and CcmF for system I and CcsA for system II).

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Cobalt-cytochrome c. I. Preparation, properties, and enzymic activity

Cited by 64 publications

References 39 publications

In Vitro Studies on Thioether Bond Formation between Hydrogenobacter thermophilus Apocytochrome c552 with Metalloprotoporphyrin Derivatives

In Vitro Studies on Thioether Bond Formation between Hydrogenobacter thermophilus Apocytochrome c552 with Metalloprotoporphyrin Derivatives

Using deeply trapped intermediates to map the cytochrome c folding landscape

Heme Concentration Dependence and Metalloporphyrin Inhibition of the System I and II CytochromecAssembly Pathways

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