The domain structure of the cholesterol side-chain cleavage cytochrome P-450 from bovine adrenocortical mitochondria

Chashchin, Vadim L.; Vasilevsky, V.I.; Shkumatov, V. M.; Ахрем, А. А.

doi:10.1016/0167-4838(84)90104-3

Cited by 26 publications

(22 citation statements)

References 30 publications

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“…To facilitate direct comparison with previous experimental data, results were normalized to 100% maximum activity corresponding to the region in the activity plot where the activity reaches a plateau. Note that the 100% value corresponds to 3.7 nmol of PG/min/nmol of P450SCC for PC (10,6) and 3.9 nmol of PG/min/nmol of P450SCC for PC (12,8), respectively. As can be seen from Fig.…”

Section: Effect Of Branched 12-diacyl Pcs On the Catalytic Activity mentioning

confidence: 99%

“…Fig. 2 shows the activity of P450SCC in vesicle preparations containing pure DMPC and mixtures of DMPC with either PC (10,6) or PC (12,8), respectively. The results demonstrate the effect of varying the membrane content of the branched 1,2-diacyl PC.…”

Section: Effect Of Branched 12-diacyl Pcs On the Catalytic Activity mentioning

confidence: 99%

“…P450SCC, adrenodoxin (AD), and adrenodoxin reductase (AR) were purified from bovine adrenocortical mitochondria to electrophoretic purity using specific affinity adsorbents (8,9). They were stored until use at Ϫ80°C in relatively high concentrated form in 50 mM phosphate buffer containing 1 M NaCl, 0.3% sodium cholate, 1 mM EDTA, and 20%(v/v) glycerol.…”

Section: Materials-[mentioning

confidence: 99%

“…For this purpose, the ␣-branched phosphatidylcholines 1,2-di-(2Ј-hexyldecanoyl)-sn-glycero-3-phosphocholine (PC (10,6)) and 1,2-di-(2Ј-octyl-dodecanoyl)-sn-glycero-phosphocholine (PC (12,8)) have been synthesized and used for reconstitution of the SCC activity of P450SCC using various phosphatidylcholine vesicle * This study was supported by Grants Schw 471/1-3 and 436 WER 17/6/94 S from the German Research Foundation (to D. S. and P. K.). The costs of publication of this article were defrayed in part by the payment of page charges.…”

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confidence: 99%

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α-Branched 1,2-Diacyl Phosphatidylcholines as Effectors of Activity of Cytochrome P450SCC (CYP11A1)

Schwarz

Kisselev

Wessel

et al. 1996

Journal of Biological Chemistry

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We synthesized phosphatidylcholines with fully saturated branched fatty acyl chains substituted in the 2-positions of the main chains and studied the influence of these lipids on the activity and other properties of P450SCC in vesicle-reconstituted systems. These saturated branched lipids, with regard to the fatty acyl moiety in molecular shape similar to cardiolipin but with the headgroup of phosphatidylcholines retained, showed a stimulatory efficiency higher than any other phospholipid and at least comparable to cardiolipin. Activation is sensitive to the acyl chain structure and composition. Results suggest that the shape of the molecule at least partially plays an important role in the process of stimulation of the activity of P450SCC. Because binding of cholesterol was increased by the branched lipids monitored optically by the fraction of P450SCC in the high spin form, it was concluded that these lipids, like cardiolipin and other lipids, exert their effects by regulating the binding of cholesterol to P450SCC. These data suggest that polymorphic lipids such as branched phosphatidylcholines and cardiolipin might influence P450SCC function by maintenance of the membrane curvature at a value optimal for activity.

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Section: Effect Of Branched 12-diacyl Pcs On the Catalytic Activity mentioning

confidence: 99%

Section: Effect Of Branched 12-diacyl Pcs On the Catalytic Activity mentioning

confidence: 99%

Section: Materials-[mentioning

confidence: 99%

mentioning

confidence: 99%

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α-Branched 1,2-Diacyl Phosphatidylcholines as Effectors of Activity of Cytochrome P450SCC (CYP11A1)

Schwarz

Kisselev

Wessel

et al. 1996

Journal of Biological Chemistry

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“…P450SCC, adrenodoxin, and adrenodoxin reductase were purified from bovine adrenocortical mitochondria to electrophoretic purity using specific affinity adsorbents (11,12). They were stored until use at Ϫ80°C in relatively high concentrated form in 50 mM phosphate buffer containing 1 M NaCl, 0.3% sodium cholate, 1 mM EDTA, and 20% (v/v) glycerol.…”

Section: Materials-[mentioning

confidence: 99%

Branched Phosphatidylcholines Stimulate Activity of Cytochrome P450SCC (CYP11A1) in Phospholipid Vesicles by Enhancing Cholesterol Binding, Membrane Incorporation, and Protein Exchange

Kisselev

Wessel

Pisch

et al. 1998

Journal of Biological Chemistry

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Phosphatidylcholines (PCs) with branched fatty acyl chains substituted in the two positions of the main chains (branched PCs) have been shown to be potent activators of the side chain cleavage activity of cytochrome P450SCC (CYP11A1) (Schwarz, D., Kisselev, P., Wessel, R., Jueptner, O., and Schmid, R. D. (1996) J. Biol. Chem. 271, 12840 -12846). The present study reports on the effect of a series of branched PC on cholesterol binding, membrane integration, and protein exchange in large unilamellar vesicles prepared by an extrusion technique. Enzyme kinetics using vesicles as well as optical titration using a micelle system with the detergent Tween 20 demonstrate that activation is correlated with the fraction of P450SCC in the high spin form. The potency of branched PCs both to activate the enzyme and to induce spin state changes increases with increasing lengths of both the branched and main fatty acyl chains. We found that the extent as well as the rate of integration of P450SCC into vesicle membranes studied by gel chromatography and stopped flow kinetics were increased by branched PC. Finally, it is demonstrated by measurement of the enzymatic activity in primary and secondary vesicles that branched PCs are potent in retaining a very rapid exchange of P450SCC between vesicles, in contrast to cardiolipin, that partially inhibits this exchange process. The data suggest that different properties of P450SCC in membrane systems including cholesterol binding, membrane integration, and protein exchange are affected by branched PCs and probably by other phospholipids, too, and therefore must be considered in an explanation of the observed high stimulation of activity.Cytochrome P450SCC (CYP11A1) is an integral mitochondrial enzyme that is located on the matrix face of the inner membrane of mitochondria. It catalyzes the side chain cleavage of cholesterol to yield pregnenolone, the common precursor of steroid hormones. Enzymatic studies employing both solution (micelle) and membrane (vesicle) systems demonstrated stimulation of activity to varying extent by phospholipids depending on the phospholipid used, the kind of reconstitution, size of vesicles, and small amounts of impurities (for a review see Ref. and references therein and Refs. 2-4). Under the lipids used in reconstitution CL1 played a special role mainly because (i) it is a typical mitochondrial lipid and (ii) it represents the most potent activator of P450SCC activity (5-7). With regard to the mechanism of activation, CL is of further interest because of the exceptional structure of its hydrophobic part, i.e. consisting of four fatty acyl chains instead of two of most other phospholipids.It has been recently reported by us that certain branched PCs that in the structure of their hydrophobic part are similar to CL but have fully saturated acyl chains cause potent activation of P450SCC comparable only with those by CL (8). By systematic variation of the lengths of both the branched and main chains of these lipids and systematic introduction of unsaturation,...

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Escherichia coliFlavodoxin Sepharose as an Affinity Resin for Cytochromes P450 and Use to Identify a Putative Cytochrome P450c17/3β-Hydroxysteroid Dehydrogenase Interaction

Jenkins

Pikuleva

Kagawa

et al. 1997

Archives of Biochemistry and Biophysics

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The domain structure of the cholesterol side-chain cleavage cytochrome P-450 from bovine adrenocortical mitochondria

Cited by 26 publications

References 30 publications

α-Branched 1,2-Diacyl Phosphatidylcholines as Effectors of Activity of Cytochrome P450SCC (CYP11A1)

α-Branched 1,2-Diacyl Phosphatidylcholines as Effectors of Activity of Cytochrome P450SCC (CYP11A1)

Branched Phosphatidylcholines Stimulate Activity of Cytochrome P450SCC (CYP11A1) in Phospholipid Vesicles by Enhancing Cholesterol Binding, Membrane Incorporation, and Protein Exchange

Escherichia coliFlavodoxin Sepharose as an Affinity Resin for Cytochromes P450 and Use to Identify a Putative Cytochrome P450c17/3β-Hydroxysteroid Dehydrogenase Interaction

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