Functional Role of Two Interhelical Disulfide Bonds in Human Cox17 Protein from a Structural Perspective

Banci, Lucia; Bertini, Ivano; Cefaro, Chiara; Ciofi‐Baffoni, Simone; Gallo, Angelo

doi:10.1074/jbc.m111.246223

Cited by 22 publications

(16 citation statements)

References 37 publications

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“…Yeast Mia40 (residues 284-403) and Cox17* C16S C23S C24S were linked N terminally with the SUMO protein Smt3 from Saccharomyces cerevisiae containing an N-terminal hexahistidine tag and cloned into the expression plasmid pET11a. For Cox17 without cysteines or with the outer or inner disulphide only, we did not routinely replace cysteines by serines, as in previous work 35 . For choosing good substitute amino acids, we considered the space occupied by the cysteines in the structure of Cox17 as well as the exposure to solvent.…”

Section: Methodsmentioning

confidence: 99%

Mia40 targets cysteines in a hydrophobic environment to direct oxidative protein folding in the mitochondria

Koch

Schmid

2014

Nat Commun

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Mia40 catalyses the oxidative folding of disulphide-containing proteins in the mitochondria. The folding pathway is directed by the formation of the first mixed disulphide between Mia40 and its substrate. Here, we employ Cox17 to elucidate the molecular determinants of this reaction. Mia40 engages initially in a dynamic non-covalent enzyme-substrate complex that forms and dissociates within milliseconds. Cys36 of Cox17 forms the mixed disulphide in an extremely rapid reaction that is limited by the preceding complex formation with Mia40. Cys36 reacts much faster than the three other cysteines of Cox17, because it neighbours three hydrophobic residues. Mia40 binds preferentially to hydrophobic regions and the dynamic nature of the non-covalent complex allows rapid reorientation for an optimal positioning of the reactive cysteine. Mia40 thus uses the unique proximity between its substrate-binding site and the catalytic disulphide to select a particular cysteine for forming the critical initial mixed disulphide.

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Section: Methodsmentioning

confidence: 99%

Mia40 targets cysteines in a hydrophobic environment to direct oxidative protein folding in the mitochondria

Koch

Schmid

2014

Nat Commun

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“…Therefore, the formation of Cu 2+ /S100A13/FGF1 ternary complex appears to be the second molecular event. The ionic form of Cu 2+ is highly toxic to the cell and therefore it is likely that the S100A13/FGF1 binary complex receives Cu 2+ from cell-surface copper chaperones, such as Atx1/CCS/Cox17 [57-61] or from SphK1, which has a very high copper affinity [11]. Cu 2+ - binding site on S100A13 and Cys 30 in FGF1 appears to be in close proximity in the Cu 2+ /S100A13/FGF1 ternary complex (Fig.7).…”

Section: Resultsmentioning

confidence: 99%

Copper binding affinity of the C2B domain of synaptotagmin-1 and its potential role in the nonclassical secretion of acidic fibroblast growth factor

Jayanthi

Kathir

Rajalingam

et al. 2014

Biochimica et Biophysica Acta (BBA) - Proteins and Proteomics

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Fibroblast growth factor 1 (FGF1) is a heparin-binding proangiogenic protein. FGF1 lacks the conventional N-terminal signal peptide required for secretion through the endoplasmic reticulum (ER) -Golgi secretory pathway. FGF1 is released through a Cu2+ - mediated nonclassical secretion pathway. The secretion of FGF1 involves the formation of a Cu2+- mediated multiprotein release complex (MRC) including FGF1, S100A13 (a calcium-binding protein) and p40 synaptotagmin (Syt1). It is believed that binding of Cu2+ to the C2B domain is important for the release of FGF1 in to the extracellular medium. In this study, using a variety of biophysical studies, Cu2+ and lipid interactions of the C2B domain of Syt1were characterized. Isothermal titration calorimetry (ITC) experiments reveal that C2B domain binds to Cu2+ in a biphasic manner involving an initial endothermic and a subsequent exothermic phase. Fluorescence energy transfer experiments using Tb3+ show that there are two Cu2+- binding pockets on the C2B domain, and one of these is also a Ca2+- binding site. Lipid-binding studies using ITC demonstrate that the C2B domain preferentially binds to small unilamellar vesicles of phosphatidyl serine (PS). Results of the differential scanning calorimetry and limited trypsin digestion experiments suggest that C2B domain is marginally destabilized upon binding to PS vesicles. These results, for the first time, suggest that the main role of the C2B domain of Syt1 is to serve as an anchor for the FGF1 MRC on the membrane bilayer. In addition, binding of the C2B domain to the lipid bilayer is shown to significantly decrease the binding affinity of the protein to Cu2+. The study provides valuable insights on the sequence of structural events that occur in the nonclassical secretion of FGF1.

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“…3). Previous studies [30,31] had shown that the inner (as opposed to the outer) disulfide of the small Tim and Cox17 substrates of Mia40 was more important for the folding and stability of these substrates. In light of these previous studies, we aimed to determine whether one of the two native disulfides of Mia40 was more important for the interaction with Erv1.…”

Section: Structural Disulfide Bonds Are Of Equal Importance For Intermentioning

confidence: 97%

Biogenesis of yeast Mia40 – uncoupling folding from import and atypical recognition features

et al. 2013

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The discovery of the mitochondrial intermembrane space assembly (MIA) pathway was followed by studies that focused mainly on the typical small substrates of this disulfide relay system and the interactions between its two central partners: the oxidoreductase Mia40 and the FAD-protein Erv1. Recent studies have revealed that more complex proteins utilize this pathway, including Mia40 itself. In the present study, we dissect the Mia40 biogenesis in distinct stages, supporting a kinetically coordinated sequence of events, starting with (a) import and insertion through the Tim23 translocon, followed by (b) folding of the core of imported Mia40 assisted by the endogenous Mia40 and (c) final interaction with Erv1. The interaction with endogenous Mia40 and the subsequent interaction with Erv1 represent kinetically distinguishable steps that rely on completely different determinants. Interaction with Mia40 proceeds very early (within 30 s) and is characterized by no Cys-specificity, an increased tolerance to mutations of the hydrophobic substrate-binding cleft and no apparent dependence on glutathione as a proofreading mechanism. All of these features illustrate a very atypical behaviour for the Mia40 precursor compared to other substrates of the MIA pathway. By contrast, interaction with Erv1 occurs after 5 min of import and relies on a more stringent specificity.

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Functional Role of Two Interhelical Disulfide Bonds in Human Cox17 Protein from a Structural Perspective

Cited by 22 publications

References 37 publications

Mia40 targets cysteines in a hydrophobic environment to direct oxidative protein folding in the mitochondria

Mia40 targets cysteines in a hydrophobic environment to direct oxidative protein folding in the mitochondria

Copper binding affinity of the C2B domain of synaptotagmin-1 and its potential role in the nonclassical secretion of acidic fibroblast growth factor

Biogenesis of yeast Mia40 – uncoupling folding from import and atypical recognition features

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