STRUCTURAL ORGANIZATION OF CYTOCHROME P-450scc FROM ADRENAL CORTEX

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

doi:10.1016/b978-0-12-187701-9.50013-9

Cited by 3 publications

(6 citation statements)

References 2 publications

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“…It should be noted that immobilized films were incubated with 50 mM TBS containing 1 M NaCl and 0.3% of sodium cholate in order to separate the covalently immobilized AD−P450scc complex and noncovalently attached AD molecules. It is well-known that under such conditions the AD and cytochrome molecules are separated and the complex cannot form . Such treated AD−P450scc monolayers were then incubated with anti-AD antibodies to determine the complex orientation.…”

Section: Resultsmentioning

confidence: 99%

“…Materials. P450scc and AD were affinity-purified using specific adsorbents from bovine adrenocortical mitochondria. , Affinity-purified polyclonal rabbit antibodies specific to AD, P450scc, and F1, F2 domains are available from previous studies. , Blocker BSA and Protein A/G were purchased from Pierce. GOPTS (reagent grade), sodium hydrogen carbonate and sodium chloride (A.C.S.…”

Section: Methodsmentioning

confidence: 99%

“…P450scc concentration was determined from CO-difference spectra (dithionite-reduced preparation) using an extinction coefficient of 91 mM -1 cm -1 for difference in the absorbance between 450 and 490 nm . AD concentration was calculated using an extinction coefficient of 9.8 mM -1 cm -1 at 414 nm . The labeling degree of Protein A/G was calculated using a molar extinction coefficient for FITC ε 493 = 62 mM -1 cm -1 in 30 mM Tris buffer, pH 7.4.…”

Section: Methodsmentioning

confidence: 99%

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Orientation of Cytochrome P450scc in Langmuir−Blodgett Monolayers

et al. 1997

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Monolayers of cytochrome P450scc and its complex with adrenodoxin were formed by Langmuir techniques and covalently immobilized on the solid substrates. The orientation of hemeprotein molecules was studied using polyclonal antibodies specific to the intact cytochrome P450scc molecule and its tryptic fragments F1 and F2, representing N- and C-terminal parts of the hemeprotein molecule. Specific interactions of the Langmuir films of cytochrome P450scc with adrenodoxin were investigated, and the position of the ferredoxin binding site at the hemeprotein molecule was identified. It was shown that the molecular orientation of the P450scc−AD (adrenodoxin) complex at the water−air interface is dependent on the surface density of the monolayer. The P450scc molecules do not denature upon spreading on the water surface. The formed monolayer can be transferred from the air−water interface to the surface modified with siloxane polymer and covalently immobilized without damage to the structure. A model considering the mode of orientation of cytochrome P450scc molecules on the air−water interface in dependence on the surface pressure is discussed.

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Section: Resultsmentioning

confidence: 99%

Section: Methodsmentioning

confidence: 99%

Section: Methodsmentioning

confidence: 99%

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Orientation of Cytochrome P450scc in Langmuir−Blodgett Monolayers

et al. 1997

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“…The crystal structure of AdR has also been solved, its size being equal to 5.8 × 5.4 × 4.0 nm [ 7 ]. As is known, the isolated membrane cytochrome CYP11A1 is able to form oligomers in solution [ 8 ]. Therefore, the structure of CYP11A1 still remains to be clarified.…”

Section: Introductionmentioning

confidence: 99%

Atomic Force Microscopy Study of Protein–Protein Interactions in the Cytochrome CYP11A1 (P450scc)-Containing Steroid Hydroxylase System

et al. 2010

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Atomic force microscopy (AFM) and photon correlation spectroscopy (PCS) were used for monitoring of the procedure for cytochrome CYP11A1 monomerization in solution without phospholipids. It was shown that the incubation of 100 μM CYP11A1 with 12% Emulgen 913 in 50 mM KP, pH 7.4, for 10 min at T = 22°C leads to dissociation of hemoprotein aggregates to monomers with the monomerization degree of (82 ± 4)%. Following the monomerization procedure, CYP11A1 remained functionally active. AFM was employed to detect and visualize the isolated proteins as well as complexes formed between the components of the cytochrome CYP11A1-dependent steroid hydroxylase system. Both Ad and AdR were present in solution as monomers. The typical heights of the monomeric AdR, Ad and CYP11A1 images were measured by AFM and were found to correspond to the sizes 1.6 ± 0.2 nm, 1.0 ± 0.2 nm and 1.8 ± 0.2 nm, respectively. The binary Ad/AdR and AdR/CYP11A1mon complexes with the heights 2.2 ± 0.2 nm and 2.8 ± 0.2 nm, respectively, were registered by use of AFM. The Ad/CYP11A1mon complex formation reaction was kinetically characterized based on optical biosensor data. In addition, the ternary AdR/Ad/CYP11A1 complexes with a typical height of 4 ± 1 nm were AFM registered.

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“…(1 99)] might derive biosynthetically from papa kusterol .4 * A recurring question concerns whether sterols of marine invertebrates are biosynthesized entirely by the animal or whether dietary sources or their biosynthesis by symbiotic algae play a major or a minor role. Zooxanthellae that were isolated from several gorgonians contained 4a-me thyl-sterols which did not possess the highly alkylated side-chains that are characteristic of the sterols of their hosts.42* On the other hand, zooxanthellae that were isolated from the sea anemone Aiptasiu pulchella synthesized both gorgosterol (205) and 23-demethylgorgosterol (206) in the absence of the Furthermore, the biosynthesis of dinosterol (172a) by the dinoflagellate Crypthecodiniurn cohnii may be relevant to the occurrence of gorgosterol in host invertebrate^.^^^ Marine phytoplankton provides potential dietary sources of steroids; in this connection, mevalonate was incorporated into cholesterol and its A22-24-methyl derivative by the phytoplankton Pseudoisochrysis p u r a d o~a . ~~~ The sponge Axinella polypoides contains a unique series of 19-nor-steroids that are typified by compounds (207a-c) and (208a-c).…”

Section: Other Invertebratesmentioning

confidence: 99%

The biosynthesis of triterpenoids and steroids

Harrison¹

1985

Nat. Prod. Rep.

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STRUCTURAL ORGANIZATION OF CYTOCHROME P-450scc FROM ADRENAL CORTEX

Cited by 3 publications

References 2 publications

Orientation of Cytochrome P450scc in Langmuir−Blodgett Monolayers

Orientation of Cytochrome P450scc in Langmuir−Blodgett Monolayers

Atomic Force Microscopy Study of Protein–Protein Interactions in the Cytochrome CYP11A1 (P450scc)-Containing Steroid Hydroxylase System

The biosynthesis of triterpenoids and steroids

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