Artículo

Estamos trabajando para incorporar este artículo al repositorio
Consulte el artículo en la página del editor
Consulte la política de Acceso Abierto del editor

Abstract:

3β-hydroxysteroid dehydrogenase 5-ene isomerase (3βHSD/I) activity is necessary for the biosynthesis of hormonally active steroids. A dual distribution of the enzyme was described in toad testes. The present study demonstrates that in testicular tissue of Bufo arenarum H., microsomal 3βHSD/I has more affinity for dehydroepiandrosterone (DHEA) than for pregnenolone (K(m) = 0.17 ± 0.03 and 1.02 μM, respectively). The Hill coefficient for the conversion of DHEA and pregnenolone were 1.04 and 1.01, respectively. The inclusion of DHEA in the kinetic analysis of pregnenolone conversion affected V(max) while K(m) was not modified, suggesting a non-competitive inhibition of the conversion of pregnenolone. K(i) was calculated from replot of Dixon's slope for each substrate concentration. K(i) from the intercept and the slope of this replot were similar (0.276 ± 0.01 and 0.263 ± 0.02 μM) and higher than the K(m) for DHEA. The K(m) and K(i) values suggest the presence of two different binding sites. When pregnenolone was present in the assays with DHEA as substrate, no effect was observed on the V(max) while K(m) values slightly increased with pregnenolone concentration. Consequently, pregnenolone inhibited the transformation of DHEA in a competitive fashion. These studies suggest that, in this species, the microsomal biosyntheses of androgens and progesterone are catalysed by different active sites. (C) 2000 Elsevier Science Ltd.

Registro:

Documento: Artículo
Título:Kinetic properties of microsomal 3β hydroxysteroid dehydrogenase-isomerase from the testis of Bufo arenarum H
Autor:Pozzi, A.G.; Canosa, L.F.; Calvo, J.C.; Ceballos, N.R.
Filiación:Programa De Regulación Hormonal Y Metabólica (PRHOM-CONICET), Buenos Aires, Argentina
Departamento De Ciencias Biológicas, Laboratorio De Endocrinología Comparada, Universidad De Buenos Aires, Buenos Aires, Argentina
Departamento De Química Biológica, Instituto De Biología Y Medicina Experimental, Universidad De Buenos Aires, Buenos Aires, Argentina
Departamento De Ciencias Biológicas, Laboratorio De Endocrinología Comparada, Ciudad Universitaria, 1428 Buenos Aires, Argentina
Palabras clave:3HSD/I; Non-competitive inhibition; Steroid biosynthesis; Toad-testis; 3(or 17)beta hydroxysteroid dehydrogenase; androstenedione; carbon 14; microsome enzyme; prasterone; pregnenolone; progesterone; steroid hormone; tritium; animal tissue; article; binding site; biosynthesis; enzyme assay; enzyme kinetics; male; microsome; nonhuman; steroid metabolism; steroidogenesis; testis; thin layer chromatography; toad
Año:2000
Volumen:73
Número:5
Página de inicio:257
Página de fin:264
DOI: http://dx.doi.org/10.1016/S0960-0760(00)00074-1
Título revista:Journal of Steroid Biochemistry and Molecular Biology
Título revista abreviado:J. Steroid Biochem. Mol. Biol.
ISSN:09600760
CODEN:JSBBE
CAS:3(or 17)beta hydroxysteroid dehydrogenase, 9015-81-0; androstenedione, 26264-53-9, 63-05-8; carbon 14, 14762-75-5; prasterone, 53-43-0; pregnenolone, 145-13-1; progesterone, 57-83-0; tritium, 10028-17-8
Registro:https://bibliotecadigital.exactas.uba.ar/collection/paper/document/paper_09600760_v73_n5_p257_Pozzi

Referencias:

  • Csapo, A.I., The 'see-saw' theory of parturition (1977), 47, pp. 159-210. , J. Knight, M. O'Connor (Eds.), The Fetus and Birth, Elsevier Amsterdam; Hall, P.F., Cellular organization for steroidogenesis (1984) Int. Rev. Cytol., 86, pp. 53-95
  • Berko, E.A., Thomas, J., Strickler, R.C., 3β-hydroxysteroid dehydrogenase in human placental microsomes and mitochondria: Co-solubilization of androstene and pregnene activities (1986) Steroids, 47, pp. 351-363
  • Thomas, J.L., Berko, E.A., Faustino, A., Myers, R.P., Strickler, R.C., Human placental 3β-hydroxy-5-ene-steroid dehydrogenase and steroid 5:4-ene-isomerase: Purification from microsomes, substrate kinetics, and inhibition by product steroids (1988) J. Steroid. Biochem., 31, pp. 785-793
  • Samuels, L.T., Helmreich, M.L., Lasuter, M.B., Reinch, H., An enzyme in endocrine tissues which oxidizes delta 5-3 hydroxyde steroids to α, β unsaturated ketones (1951) Science, 113, pp. 490-491
  • Devine, P.L., Kelly, N.S., Adams, J.B., 3β-Hydroxysteroid dehydrogenase in guinea-pig kidney: Possible involvement in 11-deoxycorticosterone formation in situ (1986) J. Steroid. Biochem., 25, pp. 265-270
  • Zhao, H.F., Rheaume, E., Trudel, J., Couet, J., Labrie, F., Simard, J., Structure and sexual dimorphic expression of a liver-specific rat 3β-hydroxysteroid dehydrogenase/isomerase (1990) Endocrinology, 127, pp. 3237-3239
  • Labrie, F., Simard, J., Luu-The, V., Belanger, A., Pelletier, G., Structure, function and tissue-specific gene expression of 3β-hydroxysteroid dehydrogenase/5-ene-4-ene isomerase enzymes in classical and peripheral intracrine steroidogenic tissues (1992) J. Steroid. Biochem. Mol. Biol., 43, pp. 805-826
  • Simard, J., Rheaume, E., Mebarki, F., Sanchez, R., New, M.I., Morel, Y., Labrie, F., Molecular basis of human 3β-hydroxysteroid dehydrogenase deficiency (1995) J. Steroid. Biochem. Mol. Biol., 53, pp. 127-138
  • Samuels, L.T., Helmrich, M.L., The influence of chorionic gonadotropin on the 3β-ol dehydrogenase of testes and adrenals (1956) Endocrinology, 58, pp. 435-442
  • Inano, H., Hayashiyama, J., Tamaoki, B.I., Solubilization of delta 5-3β-hydroxysteroid dehydrogenase with delta 5-delta 4 isomerase and 17β-hydroxysteroid dehydrogenase from rat testicular microsomal fraction by several detergents (1982) J. Steroid. Biochem., 16, pp. 587-593
  • Ishii-Ohba, H., Inano, H., Tamaoki, B.I., Purification and properties of testicular 3β-hydroxy-5-ene-steroid dehydrogenase and 5-ene-4-ene isomerase (1986) J. Steroid. Biochem., 25, pp. 555-560
  • Sulimovici, S., Bartoov, B., Lunefeld, B., Localization of 3β-hydroxysteroid dehydrogenase in the inner membrane subfraction of rat testis mitochondria (1973) Biochim. Biophys. Acta, 321, pp. 27-40
  • Pozzi, A.G., Lantos, C.P., Ceballos, N.R., Mitochondrial localization of the 3β-hydroxysteroid dehydrogenase-5-ene isomerase in interrenals of Bufo arenarum H (1996) Gen. Comp. Endocrinol., 103, pp. 176-181
  • Pozzi, A.G., Lantos, C.P., Ceballos, N.R., Subcellular localization of 3β-hydroxysteroid dehydrogenase isomerase in testis of Bufo arenarum H (1997) Gen. Comp. Endocrinol., 106, pp. 400-406
  • Townsley, J.D., Inhibition of placental 3β-hydroxysteroid dehydrogenase by naturally occurring steroids. A potential mechanism regulating oestrogen synthesis from unconjugated precursors (1975) Acta Endocrinol. (Copenhagen), 79, pp. 740-748
  • Yoshida, N., Steroid specificity of human placental 5-ene-3β-hydroxysteroid oxidoreductase (1979) Steroids, 33, pp. 9-21
  • Gibb, W., Substrate and nucleotide specificity of placental microsomal 3β-hydroxysteroid dehydrogenase (1981) Steroids, 37, pp. 23-31
  • Hall, P.F., Testicular steroid synthesis: Organization and regulation (1994), 1, pp. 1335-1362. , E. Knobil, J. Neill, (Eds.), The Physiology of Reproduction, Raven Press, New York; Botte, V., Delrio, G., Effect of estradiol-17β on the distribution of 3β-hydroxysteroid dehydrogenase in the testes of Rana esculenta and Lacerta sicula (1967) Gen. Comp. Endocrinol., 9, pp. 110-115
  • Antila, E., Saure, A., Seasonal variation of androgen interconversion in testicular tissue of Rana temporaria in vitro (1978) Experientia, 35, pp. 326-328
  • Wiebe, J.P., The mechanism of action of gonadotrophic hormones in amphibians: The stimulation of delta 5-3β-hydroxysteroid dehydrogenase activity in testes of Xenopus laevis Daudin (1970) J. Endocrinol., 47, pp. 439-450
  • Wiebe, J.P., Mechanism of gonadotropin action in Amphibia: Involvement of mitochondria (1971) Science, 175, pp. 67-68
  • Wiebe, J.P., On the mechanism and subcellular site of action of gonadotropins with respect to steroidogenic enzyme activity in testes of lower vertebrates (1972) Gen. Comp. Endocrinol., (SUPPL. 3), pp. 626-635
  • Canosa, L.F., Pozzi, A.G., Ceballos, N.R., Pregnenolone and progesterone metabolism by the testes of Bufo arenarum (1998) J. Comp. Physiol. B, 168, pp. 491-496
  • Lowry, O.H., Rosebrough, N.J., Farr, A.L., Randall, R.J., Protein measurement with the Folin phenol reagent (1951) J. Biol. Chem., 193, pp. 265-275
  • Ceballos, N.R., Cozza, E.N., Lantos, C.P., Corticoidogenesis in Bufo arenarum H. I. In vitro biosynthesis of [3H]pregnenolone and [3H]corticosterone metabolites and of endogenous 3-oxo-4-ene intermediates at 28°C and 37°C (1983) Gen. Comp. Endocrinol., 51, pp. 138-147
  • Ceballos, N.R., Shackleton, C.H., Harnik, M., Cozza, E.N., Gros, E.G., Lantos, C.P., Corticoidogenesis in the toad Bufo arenarum H: Evidence for a precursor role for an aldosterone 3β-hydroxy-5-ene analogue (3β,11β,21-trihydroxy-20-oxo-5-pregnen-18-al) (1993) Biochem. J., 292, pp. 143-147
  • Wright, R.S., A reagent for the non-destructive location of steroids and some other lipophilic materials on silica gel thin layer chromatograms (1971) J. Chromatogr., 59, pp. 220-221
  • Dixon, M., The determination of enzyme inhibitor constants (1953) Biochem. J., 137, pp. 143-144
  • Eisenthal, R., Cornish-Bowden, A., The Direct Linear Plot. A new graphical procedure for estimating enzyme kinetic parameters (1974) Biochem. J., 139, pp. 715-720
  • Lutz, R.A., Bull, C., Rodbard, D., Computer analysis of enzyme-substrate-inhibitor kinetic data with automatic model selection using IBM-PC compatible microcomputers (1986) Enzyme, 36, pp. 197-206
  • Doostzadeh, J., Morfin, R., Effects of cytochrome P450 inhibitors and of steroid hormones on the formation of 7-hydroxylated metabolites of pregnenolone in mouse brain microsomes (1997) J. Endocrinol., 155, pp. 343-350
  • Cooke, G.M., Identification of phospholipids capable of modulating the activities of some enzymes involved in androgen and 16-androstene biosynthesis in the immature pig testis (1992) J. Steroid. Biochem. Mol. Biol., 42, pp. 151-159
  • Hebert, P., Cooke, G.M., Kinetic evidence for separate 3β-hydroxysteroid dehydrogenase-isomerase in androgen and 16-androstene biosynthetic pathway in the pig testis (1992) J. Steroid. Biochem. Mol. Biol., 42, pp. 901-910
  • Perrin, A., Chambaz, E.M., Defaye, G., Modulation of hydroxylase and lyase activities of bovine cytochrome P-45017(α) in adrenal and testicular microsomes by a tissue-specific local membrane environment (1995) J. Steroid. Biochem. Mol. Biol., 54, pp. 121-129

Citas:

---------- APA ----------
Pozzi, A.G., Canosa, L.F., Calvo, J.C. & Ceballos, N.R. (2000) . Kinetic properties of microsomal 3β hydroxysteroid dehydrogenase-isomerase from the testis of Bufo arenarum H. Journal of Steroid Biochemistry and Molecular Biology, 73(5), 257-264.
http://dx.doi.org/10.1016/S0960-0760(00)00074-1
---------- CHICAGO ----------
Pozzi, A.G., Canosa, L.F., Calvo, J.C., Ceballos, N.R. "Kinetic properties of microsomal 3β hydroxysteroid dehydrogenase-isomerase from the testis of Bufo arenarum H" . Journal of Steroid Biochemistry and Molecular Biology 73, no. 5 (2000) : 257-264.
http://dx.doi.org/10.1016/S0960-0760(00)00074-1
---------- MLA ----------
Pozzi, A.G., Canosa, L.F., Calvo, J.C., Ceballos, N.R. "Kinetic properties of microsomal 3β hydroxysteroid dehydrogenase-isomerase from the testis of Bufo arenarum H" . Journal of Steroid Biochemistry and Molecular Biology, vol. 73, no. 5, 2000, pp. 257-264.
http://dx.doi.org/10.1016/S0960-0760(00)00074-1
---------- VANCOUVER ----------
Pozzi, A.G., Canosa, L.F., Calvo, J.C., Ceballos, N.R. Kinetic properties of microsomal 3β hydroxysteroid dehydrogenase-isomerase from the testis of Bufo arenarum H. J. Steroid Biochem. Mol. Biol. 2000;73(5):257-264.
http://dx.doi.org/10.1016/S0960-0760(00)00074-1