Anion Size Modulates the Structure of the A State of Cytochrome <i>c</i>

Santucci, Roberto; Bongiovanni, Cristiana; Mei, Giampiero; Ferri, Tommaso; Polizio, Francesca; Desideri, Alessandro

doi:10.1021/bi000516v

Cited by 57 publications

(88 citation statements)

References 37 publications

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“…Taken together, these data suggest that the salt opposes full protein unfolding; an equilibrium between (at least) two distinct forms, a fully unfolded high spin form and a compact non-native low spin form, is observed in solution. In agreement with previous reports, we assume the latter being the A state of cyt c, a non-native compact form with molten globule character (Goto et al 1990;Jeng et al 1990;Santucci et al 2000). The acidic transition proved to be fully reversible.…”

Section: Resultssupporting

confidence: 91%

“…5, support the absorbance data: the negatively charged 416 nmdichroic band, typical of the native protein, is almost fully recovered in bio sol-gel, while no spectral change is detected for acid-denatured cyt c embedded in pure sol-gel. On the whole, our data show that calcium nitrate plays a critical role in influencing the unfolding/ refolding process of sol-gel-embedded cyt c. In particular, the observation that the acid-denatured protein refolds only in the presence of calcium nitrate reflects the general tendency of salts to facilitate collapse of unfolded proteins into a compact state (Goto et al 1990;Jeng et al 1990;Santucci et al 2000).…”

Section: Resultsmentioning

confidence: 67%

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Spectroscopic and electrochemical characterization of cytochrome c encapsulated in a bio sol–gel matrix

et al. 2007

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Sol-gel technique represents a remarkably versatile method for protein encapsulation. To enhance sol-gel biocompatibility, systems envisaging the presence of calcium and phosphates in the sol-gel composition were recently prepared and investigated. Unfortunately, the low pH at which solutions were prepared (pH < 2.5) dramatically limited their application to proteins, because the acidic environment induces protein denaturation. In this paper we apply a new protocol based on the introduction of calcium nitrate to the inorganic phase, with formation of a binary bioactive system. In this case protein encapsulation results versatile and secure, being achieved at a pH close to neutrality (pH 6.0); also, the presence of calcium is expected to enhance system biocompatibility. To determine the properties of the salt-doped sol-gel and the influence exerted on entrapped biosystems, the structural and functional properties of embedded cytochrome c have been investigated. Data obtained indicate that the salt-doped sol-gel induces no significant change in the structure and the redox properties of the embedded protein; also, the matrix increases protein stability. Interestingly, the presence of calcium nitrate appears determinant for refolding of the acid-denatured protein. This is of interest in the perspective of future applications in biosensoristic area.

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Section: Resultssupporting

confidence: 91%

Section: Resultsmentioning

confidence: 67%

Spectroscopic and electrochemical characterization of cytochrome c encapsulated in a bio sol–gel matrix

et al. 2007

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“…The bis-His coordination of the CL-bound Lys72/73Asn mutant suggests that this conformer is less folded than the other CL-bound mutants; it is structurally similar to the molten globule state of cyt c, which also possesses a bis-His coordination. 45 Note that the far-UV CD measurements exclude any possibility of protein denaturation ( Figure S4 of the Supporting Information). The formation of a bis-His species in the double and Lys72/73Asn/His33Tyr mutants provides further evidence that Lys72 and Lys73 are crucial for protein stability.…”

Section: ■ Discussionmentioning

confidence: 99%

Role of Lysines in Cytochrome c–Cardiolipin Interaction

et al. 2013

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Cytochrome c undergoes structural variations during the apoptotic process; such changes have been related to modifications occurring in the protein when it forms a complex with cardiolipin, one of the phospholipids constituting the mitochondrial membrane. Although several studies have been performed to identify the site(s) of the protein involved in the cytochrome c−cardiolipin interaction, to date the location of this hosting region(s) remains unidentified and is a matter of debate. To gain deeper insight into the reaction mechanism, we investigate the role that the Lys72, Lys73, and Lys79 residues play in the cytochrome c−cardiolipin interaction, as these side chains appear to be critical for cytochrome c−cardiolipin recognition. The Lys72Asn, Lys73Asn, Lys79Asn, Lys72/73Asn, and Lys72/73/79Asn mutants of horse heart cytochrome c were produced and characterized by circular dichroism, ultraviolet−visible, and resonance Raman spectroscopies, and the effects of the mutations on the interaction of the variants with cardiolipin have been investigated. The mutants are characterized by a subpopulation with non-native axial coordination and are less stable than the wild-type protein. Furthermore, the mutants lacking Lys72 and/or Lys79 do not bind cardiolipin, and those lacking Lys73, although they form a complex with the phospholipid, do not show any peroxidase activity. These observations indicate that the Lys72, Lys73, and Lys79 residues stabilize the native axial Met80−Fe(III) coordination as well as the tertiary structure of cytochrome c. Moreover, while Lys72 and Lys79 are critical for cytochrome c−cardiolipin recognition, the simultaneous presence of Lys72, Lys73, and Lys79 is necessary for the peroxidase activity of cardiolipin-bound cytochrome c.

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“…Before measurements, the solution was deaerated for 30 min by a gentle flow of pure nitrogen maintained just above the solution surface. Further measurements were performed using a procedure for the chemical modification of the gold electrode as previously described [9]. Briefly, the electrode was dipped in a saturated aqueous (6-mercaptopurine) solution for 15 min.…”

Section: Electrochemical Measurementsmentioning

confidence: 99%

Conversion of cytochrome c into a peroxidase: Inhibitory mechanisms and implication for neurodegenerative diseases

Patriarca

Polticelli

Piro

et al. 2012

Archives of Biochemistry and Biophysics

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a b s t r a c tA further function of cytochrome c (cyt c), beyond respiration, is realized outside mitochondria in the apoptotic program. In the early events of apoptosis, the interaction of cyt c with a mitochondrion-specific phospholipid, cardiolipin (CL), brings about a conformational transition of the protein and acquirement of peroxidase activity. The hallmark of cyt c with peroxidase activity is its partial unfolding accompanied by loosening of the Fe sixth axial bond and an enhanced access of the heme catalytic site to small molecules like H 2 O 2 . To investigate the peroxidase activity of non-native cyt c, different forms of the protein were analyzed with the aim to correlate their structural features with the acquired enzymatic activity and apoptogenic properties (wt cyt c/CL complex and two single cyt c variants, H26Y and Y67H, free and bound to CL). The results suggest that cyt c may respond to different environments by changing its fold thus favouring the exertion of different biological functions in different pathophysiological cell conditions. Transitions among different conformations are regulated by endogenous molecules such as ATP and may be affected by synthetic molecules such as minocycline, thus suggesting a mechanism explaining its use as therapeutic agent impacting on disease-associated oxidative and apoptotic mechanisms.

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Anion Size Modulates the Structure of the A State of Cytochrome c

Cited by 57 publications

References 37 publications

Spectroscopic and electrochemical characterization of cytochrome c encapsulated in a bio sol–gel matrix

Spectroscopic and electrochemical characterization of cytochrome c encapsulated in a bio sol–gel matrix

Role of Lysines in Cytochrome c–Cardiolipin Interaction

Conversion of cytochrome c into a peroxidase: Inhibitory mechanisms and implication for neurodegenerative diseases

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