Study of the Hansenula anomala yeast flavocytochrome‐b2‐cytochrome‐c complex

Thomas, Marie‐Antoinette; Gervais, Michel; Favaudon, Vincent; Valat, P.

doi:10.1111/j.1432-1033.1983.tb07691.x

Cited by 22 publications

(11 citation statements)

References 16 publications

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“…This is much shorter than the 3.2 ns reported by Vanderkooi et al [19]. On the other hand Thomas et al [36] have reported a lifetime of 2.0 ns of the Zn-substituted derivative of Hansenula anomala cytochrome c at 10°C, which is close to our value.…”

Section: Time-resolvedfluoresence Of the Proteins In Aqueous Solutionsupporting

confidence: 38%

Time-resolved fluorescence and circular dichroism of porphyrin cytochrome c and Zn-porphyrin cytochrome c incorporated in reversed micelles

Vos¹,

Laane²,

Weijers³

et al. 1987

Eur J Biochem

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Interactions between fluorescent horse heart cytochrome c derivatives (e. g. porphyrin cytochrome c and Znporphyrin cytochrome c) with surfactant interfaces in reversed micellar solutions have been studied, using different spectroscopic techniques. Anionic [sodium bis(2-ethylhexyl)sulfosuccinate, AOT] and cationic (cetyltrimethylammonium bromide, CTAB) surfactant solutions have been used in order to investigate the effects of charge interactions between proteins and interfaces.Circular dichroism reveals that much of the protein secondary structure is lost in AOT-reversed micelles, especially when the molar water/surfactant ratio, wo, is high (w, = 40), whereas in CTAB-reversed micelles secondary structure seems to be preserved.Time-resolved fluorescence measurements of the porphyrin in the cytochrome c molecule yields information about the changes in structure and the dynamics of the protein upon interaction with surfactant assemblies both in aqueous and in hydrocarbon solutions. With AOT as surfactant a strong interaction between protein and interface can be observed. The effects found in aqueous AOT solution are of the same kind as in hydrocarbon solution. In the CTAB systems the interactions between protein and surfactant are much less pronounced. The measured effects on the fluorescence properties of the proteins are different in aqueous and hydrocarbon solutions.In general, the observations can be explained by an electrostatic attraction between the overall positively charged protein molecules and the anionic AOT interface. Electrostatic attraction can also occur between the cytochrome c derivatives and CTAB because there is a negatively charged zone on the surface of the proteins. From the fluorescence anisotropy decays it can be concluded that in the CTAB-reversed micellar system these interactions are not important, whereas in an aqueous CTAB solution the proteins interact with surfactant molecules.Surfactant assemblies in organic media, called reversed micelles or, more general, water-in-oil microemulsions, have been investigated extensively over the past years. Many studies have been carried out towards the elucidation of structure and dynamics of reversed micelles [I -41 but also on the (bio)chemical and (bio)technological applications of these systems [2, 3, 5-91. In our laboratory an important line of research consists of the incorporation of proteins in reversed micelles in order to perform bioconversions of apolar compounds or to isolate proteins [lo -121. At present, the effects of protein solubilization on both protein and micelle structure and dynamics are not very clear. Because reversed micellar solutions are optically transparent, spectroscopic and ultracentrifugation techniques can be applied to study these effects. Several systems have been the subject of this kind of investigation, some containing cytoplasmatic, others containing membrane proteins [5, 7-91. Horse heart cytochrome c is a protein which in vivo is strongly associated via electrostatic interactions with the inner Correspo...

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Section: Time-resolvedfluoresence Of the Proteins In Aqueous Solutionsupporting

confidence: 38%

Time-resolved fluorescence and circular dichroism of porphyrin cytochrome c and Zn-porphyrin cytochrome c incorporated in reversed micelles

Vos¹,

Laane²,

Weijers³

et al. 1987

Eur J Biochem

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“…Fluorescence studies of the interaction between zincsubstituted cytochrome c and flavocytochrome b2 from Hansenula anomala suggest that major portion of the binding site for cytochrome c involves the surface of the flavinbinding domain (36). These studies also indicate relatively low affinity of cytochrome c for the cytochrome b2 core.…”

mentioning

confidence: 90%

Three-dimensional structure of flavocytochrome b2 from baker's yeast at 3.0-A resolution.

Xia¹,

Shamala²,

Bethge³

et al. 1987

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

120

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“…The cytochrome b2 core and the flavodehydrogenase can be isolated after cleavage of flavocytochrome b2 by controlled proteolysis (Gervais et al, 1977). Fluorescence studies demonstrated that cytochrome c can bind both to the flavodehydrogenase (Thomas et al, 1983) and to the cytochrome b2 domain (Thomas et al, 1983; Albani, 1985), the affinity to the flavodehydrogenase domain (Kd = 1?-7 M) being higher than that to the cytochrome b2 (Kd = 10-6 M). In both cases the stoichiometry was 1: 1, as previously demonstrated for the flavocytochrome b2-cytochrome c complex (Prats, 1977).…”

Section: Introductionmentioning

confidence: 99%

Cytochrome b2, an electron carrier between flavocytochrome b2 and cytochrome c. Rapid kinetic characterization of the electron-transfer parameters with ionic-strength-dependence

Capeillère‐Blandin

Albani

1987

Biochemical Journal

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The oxidation-reduction properties of free cytochrome b2 isolated by controlled proteolysis from flavocytochrome b2, i.e. the flavodehydrogenase-bound cytochrome b2, were investigated by using stopped-flow spectrophotometry. The rapid kinetics of the reduction of cytochrome b2 by flavocytochrome b2 in the presence of L-lactate are reported. The self-exchange rate constant between reduced cytochrome b2 bound to the flavodehydrogenase and free cytochrome b2 was determined to be 10(5) M-1 X S-1 at 5 degrees C, I 0.2 and pH 7.0. The specific electron-transfer reaction between reduced cytochrome b2 and cytochrome c was also studied, giving an apparent second-order rate constant of 10(7) M-1 X S-1 at 5 degrees C, I 0.2 and pH 7.0. This electron-exchange rate is slightly modulated by ionic strength, following the Debye-Hückel relationship with a charge factor Z1Z2 = -1.9. Comparison of these data with those for the reduction of cytochrome c by flavodehydrogenase-bound cytochrome b2 [Capeillère-Blandin (1982) Eur. J. Biochem. 128, 533-542] leads to the conclusion that the intramolecular electron exchange between haem b2 and haem c within the reaction complex occurs at a rate very similar to that determined experimentally in presence of the flavodehydrogenase domain. The low reaction rate observed with free cytochrome b2 is ascribed to the low stability of the reaction complex formed between free cytochrome b2 and cytochrome c.

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Study of the Hansenula anomala yeast flavocytochrome‐b₂‐cytochrome‐c complex

Cited by 22 publications

References 16 publications

Time-resolved fluorescence and circular dichroism of porphyrin cytochrome c and Zn-porphyrin cytochrome c incorporated in reversed micelles

Time-resolved fluorescence and circular dichroism of porphyrin cytochrome c and Zn-porphyrin cytochrome c incorporated in reversed micelles

Three-dimensional structure of flavocytochrome b2 from baker's yeast at 3.0-A resolution.

Cytochrome b2, an electron carrier between flavocytochrome b2 and cytochrome c. Rapid kinetic characterization of the electron-transfer parameters with ionic-strength-dependence

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