Probing the cytochrome c′ folding landscape

Pletneva, Ekaterina V.; Zhao, Ziqing; Kimura, Tetsunari; Petrova, Krastina V.; Gray, Harry B.; Winkler, Jay R.

doi:10.1016/j.jinorgbio.2007.06.019

Cited by 10 publications

(7 citation statements)

References 35 publications

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“…The discrimination in structural behaviour suggested by the above calculations is consistent with one of the conclusions reached previously [10]. Specifically, there is no reason to suppose that an overall folding or unfolding process can be described by a two-state model.…”

Section: Molecular Physics 913supporting

confidence: 89%

“…This scenario is unlikely for the ferric protein, since the affinity of methionine for an Fe(III) heme is low in the denatured state [9]. A study of the folding kinetics of c 0 (monitored by heme absorption and native Trp72 fluorescence at pH 5) suggests deviation from two-state behaviour that features compaction of the polypeptide ensemble [10] as well as a prominent slow phase.…”

Section: Introductionmentioning

confidence: 99%

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Unfolding four-helix bundles

Gray

Kozak

2011

Molecular Physics

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A geometrical model has been developed to describe the early stages of unfolding of cytochromes c 0 and c-b 562 . Calculations are based on a step-wise extension of the polypeptide chain subject to the constraint that the spatial relationship among the residues of each triplet is fixed by the native-state crystallographic data. The response of each protein to these structural perturbations allows the evolution of each of the four helices in these two proteins to be differentiated. It is found that the two external helices in c 0 unfold before its two internal helices, whereas exactly the opposite behaviour is demonstrated by c-b 562 . Each of these cytochromes has an extended, internal, non-helical ('turning') region that initially lags behind the most labile helix but then, at a certain stage (identified for each cytochrome), unravels before any of the four helices present in the native structure. It is believed that these predictions will be useful in guiding future experimental studies on the unfolding of these two cytochromes.

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Section: Molecular Physics 913supporting

confidence: 89%

Section: Introductionmentioning

confidence: 99%

Unfolding four-helix bundles

Gray

Kozak

2011

Molecular Physics

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“…Moreover, as elaborated in our earlier studies [2][3][4], the model accounts for experimental evidence on cytochromes c [10], c [11], and cb 562 [12]. However, spatial correlations are only part of the picture.…”

Section: Introductionmentioning

confidence: 85%

Euclidean perspective on the unfolding of azurin: angular correlations

et al. 2013

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The geometrical model introduced previously by the authors has been extended quantitatively to document changes in angular correlations between and among residues as azurin unfolds. In the early stages of denaturation, these changes are found to be more pronounced than changes in the spatial displacement of residues, a result that is also found for residues acting in concert, viz., α-helices, β-sheet residues and residues in 'turning regions.' Our analysis leads to a picture of the large-scale motion of the polypeptide chain as azurin denatures. Flanking a central 'ribbon' of residues whose orientation remains essentially invariant, we find that in the early stages of unfolding, left-and right-hand 'wings' adjacent to this stationary scaffolding pivot counterclockwise, while smaller regions on opposing ends of the β-barrel pivot clockwise. As spatial constraints characterising the native state are further relaxed, our calculations show that some regions reverse their orientational motion, reflecting the enhanced flexibility of the polypeptide chain in the denatured state.

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“…We have examined a family of four-helix bundle cytochromes [cyt b 562 (24), cyt cb 562 (25,26), cyt c 556 (25), and cyt cЈ (27)(28)(29)(30)] in which refolding times differ by many orders of magnitude despite their strongly conserved structural topology (Ϸ3 Å rmsd) (25,28,(31)(32)(33). In the b-type cytochromes (e.g., cyt b 562 ), the heme is attached to the polypeptide only through axial Fe ligation.…”

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confidence: 99%

Folding energy landscape of cytochrome cb ₅₆₂

Kimura

Lee

Gray

et al. 2009

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

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Cytochrome cb562 is a variant of an Escherichia coli four-helix bundle b-type heme protein in which the porphyrin prosthetic group is covalently ligated to the polypeptide near the terminus of helix 4. Studies from other laboratories have shown that the apoprotein folds rapidly without the formation of intermediates, whereas the holoprotein loses heme before native structure can be attained. Time-resolved fluorescence energy transfer (TRFET) measurements of cytochrome cb562 refolding triggered using an ultrafast continuous-flow mixer (150 s dead time) reveal that heme attachment to the polypeptide does not interfere with rapid formation of the native structure. Analyses of the TRFET data produce distributions of Trp-59 -heme distances in the protein before, during, and after refolding. Characterization of the moments and time evolution of these distributions provides compelling evidence for a refolding mechanism that does not involve significant populations of intermediates. These observations suggest that the cytochrome b562 folding energy landscape is minimally frustrated and able to tolerate the introduction of substantial perturbations (i.e., the heme prosthetic group) without the formation of deep misfolded traps.four-helix bundle ͉ minimal frustration ͉ protein folding ͉ time-resolved fluorescence energy transfer ͉ tryptophan E nergy landscape theory has delineated principles that underlie the conversion of disordered polypeptides into correctly folded functional proteins (1-8). A key element of this theory is the concept of minimal frustration that, in its qualitative formulation, predicts that the folding energy landscape is funneled toward the native structure and does not contain a large number of deep misfolded traps. This notion derives in part from the many experimental observations of proteins that rapidly fold to native structures without the apparent population of intermediates. Additional features of minimal frustration are the robustness of protein structures to mutation and the malleability of folding pathways (1, 3-5, 7).The incorporation of prosthetic groups into protein structures introduces additional challenges for understanding folding and the maintenance of minimally frustrated folding pathways. The heme cofactors in b-and c-type cytochromes are a case in point. The apoprotein of mitochondrial cytochrome c (cyt c) is unstructured, demonstrating that heme is required to stabilize the native fold (9). The covalently bound porphyrin plays an integral role in cyt c folding, possibly as a hydrophobic nucleation site, but does not lead to nonnative clusters and misfolded traps (10). Many b-type cytochromes (e.g., cyt b 5 and cyt b 562 ), however, adopt native or near-native structures in the absence of their noncovalently bound porphyrins (11-23). On the basis of these observations, it has been suggested that folding precedes heme incorporation in the b-type cytochromes, whereas heme attachment is a prerequisite for folding in the c-type proteins. Because b-type cytochromes likely evolved to ...

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Probing the cytochrome c′ folding landscape

Cited by 10 publications

References 35 publications

Unfolding four-helix bundles

Unfolding four-helix bundles

Euclidean perspective on the unfolding of azurin: angular correlations

Folding energy landscape of cytochrome cb ₅₆₂

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Probing the cytochrome c′ folding landscape

Cited by 10 publications

References 35 publications

Unfolding four-helix bundles

Unfolding four-helix bundles

Euclidean perspective on the unfolding of azurin: angular correlations

Folding energy landscape of cytochrome cb 562

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Product

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Folding energy landscape of cytochrome cb ₅₆₂