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. 1988 Nov 15;256(1):53–59. doi: 10.1042/bj2560053

Specific accumulation of 17 alpha-hydroxyprogesterone in microsomal membranes during the process of cytochrome P-450(C-17)-catalysed androgen biosynthesis. A dynamic study of intermediate formation and turnover.

N Kühn-Velten 1, M Lessmann 1, M E Förster 1, W Staib 1
PMCID: PMC1135367  PMID: 3223911

Abstract

A complete dynamic analysis of cytochrome P-450(C-17)-catalysed androgen biosynthesis from a single dose of progesterone and 17 alpha-hydroxyprogesterone in a double-label double-substrate experiment was performed in order to elucidate the controversial intermediacy of 17 alpha-hydroxyprogesterone. Label distribution within the steroid fractions as well as in the membrane and buffer compartments yields direct evidence that the endogenously formed 17 alpha-hydroxyprogesterone (which is in an 'intermediate state') accumulates to a higher degree in microsomal membranes than does the exogenously added 17 alpha-hydroxyprogesterone (which is in a 'substrate state') under certain conditions. It is also demonstrated that endogenously formed 17 alpha-hydroxyprogesterone may partly leave the membrane compartment (in terms of a 'leakage' or 'overflow' phenomenon) and is then able to equilibrate with the pool of exogenously added 17 alpha-hydroxyprogesterone. Since only the label distribution in the membrane-associated (but not always in the aqueous) 17 alpha-hydroxyprogesterone pool corresponds to the label distribution in the androgen fraction, it is concluded that only the membrane-associated 17 alpha-hydroxyprogesterone pool is directly accessible to cytochrome P-450(C-17)-catalysed conversion into androgens.

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Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. Becker S., Chubb C., Ewing L. Mathematical model of steroidogenesis in rat and rabbit testes. Am J Physiol. 1980 Jul;239(1):R184–R195. doi: 10.1152/ajpregu.1980.239.1.R184. [DOI] [PubMed] [Google Scholar]
  2. Chasalow F. Mechanism and control of rat testicular steroid synthesis. J Biol Chem. 1979 Apr 25;254(8):3000–3005. [PubMed] [Google Scholar]
  3. Cheng B., Hsu D. K., Kimura T. Utilization of intramitochondrial membrane cholesterol by cytochrome P-450-dependent cholesterol side-chain cleavage reaction in bovine adrenocortical mitochondria: steroidogenic and non-steroidogenic pools of cholesterol in the mitochondrial inner membranes. Mol Cell Endocrinol. 1985 May;40(2-3):233–243. doi: 10.1016/0303-7207(85)90179-0. [DOI] [PubMed] [Google Scholar]
  4. Clegg J. S. Properties and metabolism of the aqueous cytoplasm and its boundaries. Am J Physiol. 1984 Feb;246(2 Pt 2):R133–R151. doi: 10.1152/ajpregu.1984.246.2.R133. [DOI] [PubMed] [Google Scholar]
  5. Eckstein B., Borut A., Cohen S. Production of testosterone from progesterone by rat testicular microsomes without release of the intermediates 17 alpha-hydroxyprogesterone and androstenedione. Eur J Biochem. 1987 Jul 15;166(2):425–429. doi: 10.1111/j.1432-1033.1987.tb13533.x. [DOI] [PubMed] [Google Scholar]
  6. Estabrook R. W., Martinez-Zedillo G., Young S., Peterson J. A., McCarthy J. The interaction of steroids with liver microsomal cytochrome P-450--a general hypothesis. J Steroid Biochem. 1975 Mar-Apr;6(3-4):419–425. doi: 10.1016/0022-4731(75)90166-1. [DOI] [PubMed] [Google Scholar]
  7. FOLCH J., LEES M., SLOANE STANLEY G. H. A simple method for the isolation and purification of total lipides from animal tissues. J Biol Chem. 1957 May;226(1):497–509. [PubMed] [Google Scholar]
  8. Fevold H. R. Regulation of the adrenal and gonadal microsomal mixed function oxygenases of steroid hormone biosynthesis. Annu Rev Physiol. 1983;45:19–36. doi: 10.1146/annurev.ph.45.030183.000315. [DOI] [PubMed] [Google Scholar]
  9. Hall P. F. Cytochromes P-450 and the regulation of steroid synthesis. Steroids. 1986 Sep-Oct;48(3-4):131–196. doi: 10.1016/0039-128x(86)90002-4. [DOI] [PubMed] [Google Scholar]
  10. Hochberg R. B., McDonald P. D., Ladany S., Lieberman S. Transient intermediates in steroidogenesis. J Steroid Biochem. 1975 Mar-Apr;6(3-4):323–327. doi: 10.1016/0022-4731(75)90150-8. [DOI] [PubMed] [Google Scholar]
  11. Hume R., Kelly R. W., Taylor P. L., Boyd G. S. The catalytic cycle of cytochrome P-450scc and intermediates in the conversion of cholesterol to pregnenolone. Eur J Biochem. 1984 May 2;140(3):583–591. doi: 10.1111/j.1432-1033.1984.tb08142.x. [DOI] [PubMed] [Google Scholar]
  12. Jost P., Griffith O. H., Capaldi R. A., Vanderkooi G. Identification and extent of fluid bilayer regions in membranous cytochrome oxidase. Biochim Biophys Acta. 1973 Jun 22;311(2):141–152. doi: 10.1016/0005-2736(73)90261-7. [DOI] [PubMed] [Google Scholar]
  13. Kominami S., Itoh Y., Takemori S. Studies on the interaction of steroid substrates with adrenal microsomal cytochrome P-450 (P-450C21) in liposome membranes. J Biol Chem. 1986 Feb 15;261(5):2077–2083. [PubMed] [Google Scholar]
  14. Krainev A. G., Weiner L. M., Alferyev I. S., Slynko N. M. Bifunctional compound study of the active-centre location of cytochrome P-450 in a microsomal membrane ('float' molecules method). Biochim Biophys Acta. 1985 Aug 8;818(1):96–104. doi: 10.1016/0005-2736(85)90143-9. [DOI] [PubMed] [Google Scholar]
  15. Kühn-Velten N., Bos D., Staib W. Differential down-regulation and induction responses of testicular steroidogenic cytochromes P-450(cscc) and P-450(C17 alpha) to human choriogonadotropin. Biosci Rep. 1986 May;6(5):451–457. doi: 10.1007/BF01116136. [DOI] [PubMed] [Google Scholar]
  16. Kühn-Velten N., Staib W. Distribution of progesterone-binding cytochrome P450 and steroid-17 alpha-hydroxylase/C-17,20-lyase within different compartments of the rat testis. FEBS Lett. 1983 Apr 5;154(1):70–74. doi: 10.1016/0014-5793(83)80877-1. [DOI] [PubMed] [Google Scholar]
  17. Kühn-Velten N., Staib W. Product inhibition of steroid-17 alpha-monooxygenase by endogenous 17 alpha-hydroxyprogesterone in microsomes and isolated Leydig cells from rat testis. J Steroid Biochem. 1984 Dec;21(6):697–700. doi: 10.1016/0022-4731(84)90033-5. [DOI] [PubMed] [Google Scholar]
  18. Kühn-Velten N., Wolff J., Staib W. Estimation of kinetic parameters of androgen-synthesizing enzyme activities in superfused Leydig cells from rat testes: difference between endogenous and exogenous substrates. Biosci Rep. 1984 Jun;4(6):483–488. doi: 10.1007/BF01122223. [DOI] [PubMed] [Google Scholar]
  19. Markwell M. A., Haas S. M., Tolbert N. E., Bieber L. L. Protein determination in membrane and lipoprotein samples: manual and automated procedures. Methods Enzymol. 1981;72:296–303. doi: 10.1016/s0076-6879(81)72018-4. [DOI] [PubMed] [Google Scholar]
  20. McCarthy J. L., Waterman M. R. Co-induction of 17 alpha-hydroxylase and C-17,20-lyase activities in primary cultures of bovine adrenocortical cells in response to ACTH treatment. J Steroid Biochem. 1988 Mar;29(3):307–312. doi: 10.1016/0022-4731(88)90031-3. [DOI] [PubMed] [Google Scholar]
  21. Menard R. H., Purvis J. L. Studies of cytochrome P-450 in testis microsomes. Arch Biochem Biophys. 1973 Jan;154(1):8–18. doi: 10.1016/0003-9861(73)90029-5. [DOI] [PubMed] [Google Scholar]
  22. Nakajin S., Hall P. F., Onoda M. Testicular microsomal cytochrome P-450 for C21 steroid side chain cleavage. Spectral and binding studies. J Biol Chem. 1981 Jun 25;256(12):6134–6139. [PubMed] [Google Scholar]
  23. Nakajin S., Hall P. F. Side-chain cleavage of C21 steroids to C19 steroids by testicular microsomal cytochrome P-450: 17alpha-hydroxy C21 steroids as obligatory intermediates. J Steroid Biochem. 1981 Dec;14(12):1249–1252. doi: 10.1016/0022-4731(81)90328-9. [DOI] [PubMed] [Google Scholar]
  24. OMURA T., SATO R. THE CARBON MONOXIDE-BINDING PIGMENT OF LIVER MICROSOMES. I. EVIDENCE FOR ITS HEMOPROTEIN NATURE. J Biol Chem. 1964 Jul;239:2370–2378. [PubMed] [Google Scholar]
  25. Samuels L. T., Bussmann L., Matsumoto K., Huseby R. A. Organization of androgen biosynthesis in the testis. J Steroid Biochem. 1975 Mar-Apr;6(3-4):291–296. doi: 10.1016/0022-4731(75)90145-4. [DOI] [PubMed] [Google Scholar]
  26. Samuels L. T., Matsumoto K. Localization of enzymes involved in testosterone biosynthesis by the mouse testis. Endocrinology. 1974 Jan;94(1):55–60. doi: 10.1210/endo-94-1-55. [DOI] [PubMed] [Google Scholar]
  27. Seybert D. W., Lancaster J. R., Jr, Lambeth J. D., Kamin H. Participation of the membrane in the side chain cleavage of cholesterol. Reconstitution of cytochrome P-450scc into phospholipid vesicles. J Biol Chem. 1979 Dec 10;254(23):12088–12098. [PubMed] [Google Scholar]
  28. Sheets J. J., Estabrook R. W. Multiple sites of steroid hydroxylation by the liver microsomal cytochrome P-450 system: primary and secondary metabolism of androstenedione. Biochemistry. 1985 Nov 5;24(23):6591–6597. doi: 10.1021/bi00344a043. [DOI] [PubMed] [Google Scholar]
  29. Suhara K., Fujimura Y., Shiroo M., Katagiri M. Multiple catalytic properties of the purified and reconstituted cytochrome P-450 (P-450sccII) system of pig testis microsomes. J Biol Chem. 1984 Jul 25;259(14):8729–8736. [PubMed] [Google Scholar]
  30. Welch G. R., Keleti T., Vértessy B. The control of cell metabolism for homogeneous vs. heterogeneous enzyme systems. J Theor Biol. 1988 Feb 21;130(4):407–422. doi: 10.1016/s0022-5193(88)80206-6. [DOI] [PubMed] [Google Scholar]
  31. Welch G. R., Somogyi B., Matkó J., Papp S. Effect of viscosity on enzyme-ligand dissociation. II. Role of the microenvironment. J Theor Biol. 1983 Jan 21;100(2):211–238. doi: 10.1016/0022-5193(83)90348-x. [DOI] [PubMed] [Google Scholar]
  32. Yoshida K., Takahashi J., Winters S. J., Oshima H., Troen P. Steroidogenesis in the monkey testis: relationship of enzyme organization to endogenous steroids, steroidogenesis and gonadotropin treatment. J Steroid Biochem. 1984 Jul;21(1):49–58. doi: 10.1016/0022-4731(84)90059-1. [DOI] [PubMed] [Google Scholar]

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