Studies of the guinea pig adrenal cytochrome P450c17 cDNA

Tremblay, Yves; Bélanger, Alain; Fleury, Alain; Beaudoin, Claude; Provost, Pierre R.; Martineau, Isabelle

doi:10.3109/07435809509030467

Cited by 9 publications

(9 citation statements)

References 16 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…This result differs from the earlier report (Tremblay et al, 1995) using COS-1 cells transfected with the cDNA for guinea pig P450c17 where P4 was reported to be 17α-hydroxylated more rapidly than P5. In addition, there was the unexplained difference that a small but measurable amount of DHEA is formed from P5 using the transfected COS-1 cell system (Tremblay et al, 1995). No DHEA was detected in the present study using the purified fusion protein assay in the presence or absence of added b 5 .…”

Section: P4 and P5 Metabolism By The Guinea Pig 17a-hydroxylase Fusiocontrasting

confidence: 85%

Comparison of the 17α-Hydroxylase/C17,20-Lyase Activities of Porcine, Guinea Pig and Bovine P450c17 Using Purified Recombinant Fusion Proteins Containing P450c17 Linked to NADPH-P450 Reductase

Shet

Fisher

Tremblay

et al. 2007

Drug Metabolism Reviews

Self Cite

View full text Add to dashboard Cite

The cDNAs for cytochrome P450c17 (P450c17) of three species, pig, guinea pig, and cow, representing three families of mammals (suidae, procaviidae, and bovidae, respectively) were each engineered into an expression plasmid (pCWori+). The P450c17 domain of the coding sequence was connected to a truncated form of rat NADPH-P450 reductase by a linker sequence encoding two amino acids (SerThr). These fusion proteins were expressed in E. coli and purified for use in enzymatic assays to determine similarities and differences in 17 alpha-hydroxylase and lyase activities. The fusion proteins were found to catalyze both the 17 alpha-hydroxylation of progesterone (P4) and pregnenolone (P5) to 17 alpha-hydroxylated P4 and P5 (17 alpha-OH P4 and 17 alpha-OH P5) followed by the C17,20-lyase reaction for the conversion of these C(21)-17 alpha-hydroxylated steroids to C(19)-steroids (the C17,20-lyase reaction). These in vitro studies show that (a) porcine P450c17 possesses cytochrome b(5) (b(5))-stimulated C17,20-lyase activity that converts 17 alpha OH-P4 to androstenedione (AD) but also converts 17 alpha-OHP5 to dehydroepiandrosterone (DHEA); (b) guinea pig P450c17 possesses a b(5)-stimulated C17,20-lyase activity that converts 17 alpha-OH P4 to AD but does not convert 17 alpha-OH P5 to DHEA., and (c) bovine P450c17 possesses a b(5)-stimulated C17,20-lyase activity that converts 17 alpha-OH P5 to DHEA but does not convert 17 alpha-OH P4 to AD. Thus, the P450c17 of each species differs in its ability to catalyze in vitro the conversion of C(21)-steroids to C(19)-steroids. In addition, each P450c17 is capable of catalyzing additional hydroxylation reactions leading to low levels of 2 alpha-, 6 beta-, 16- and 21-hydroxy-metabolites. Porcine P450c17 also catalyzes the b(5)-dependent synthesis of andien-beta (androsta-5,16-dien-3beta-ol) from P5. When the amino acid sequences of the three P450c17s were aligned there was an approximate 50% variation in the alignment identity (227 differences in the sequences of 509 amino acids). Alignment did not permit the assignment of specific amino acids or domains to the observed differences in enzymatic activities.

show abstract

Section: P4 and P5 Metabolism By The Guinea Pig 17a-hydroxylase Fusiocontrasting

confidence: 85%

Comparison of the 17α-Hydroxylase/C17,20-Lyase Activities of Porcine, Guinea Pig and Bovine P450c17 Using Purified Recombinant Fusion Proteins Containing P450c17 Linked to NADPH-P450 Reductase

Shet

Fisher

Tremblay

et al. 2007

Drug Metabolism Reviews

Self Cite

View full text Add to dashboard Cite

show abstract

“…Furthermore, even in the presence of cytochrome b 5 , the rate of the 17,20-lyase activity remains lower than that of the hydroxylase activity and is usually preferentially enhanced in either the ⌬4 or the ⌬5 pathway. For instance, in small animals, such as rats (21), mice (22), hamsters (23), and guinea pigs (24), CYP17 is more active in the ⌬4 pathway. In contrast, human and primate CYP17s utilize the ⌬5 pathway (10,11).…”

Section: Discussionmentioning

confidence: 99%

Xenopus laevis CYP17 Regulates Androgen Biosynthesis Independent of the Cofactor Cytochrome b5

Yang

Hammes

2005

Journal of Biological Chemistry

View full text Add to dashboard Cite

The enzyme CYP17 primarily regulates androgen production by mediating four reactions: conversion of pregnenolone and progesterone to 17-hydroxypregnenolone and 17-hydroxyprogesterone, respectively (17␣-hydroxylase activity), followed by conversion of the 17-hydroxylated steroids to dehydroepiandrosterone and androstenedione, respectively (17,20-lyase activity). Most mammalian CYP17 isoforms have high 17␣-hydroxylase relative to 17,20-lyase activities and preferentially mediate one of the two 17,20-lyase reactions. In contrast, Xenopus laevis CYP17 potently regulates all four reactions in the frog ovary. CYP17 isoforms generally rely on the cofactor cytochrome b 5 for the 17,20-lyase reaction, suggesting that the high lyase activity of Xenopus CYP17 might be due to a lesser dependence on b 5 . The kinetics of Xenopus CYP17 expressed in yeast microsomes were therefore examined in the absence and presence of Xenopus on human b 5 . Xenopus CYP17 mediated both 17,20-lyase reactions in the absence of b 5 , confirming that the activity did not require b 5 . However, both Xenopus and human b 5 slightly enhanced Xenopus CYP17-mediated lyase activity, indicating that the enzyme was still at least partially responsive to b 5 . Surprisingly, only the human b 5 cofactor enhanced human CYP17-mediated lyase activity, implying that the human enzyme had more specific cofactor requirements than Xenopus CYP17. Studies using human/Xenopus chimeric b 5 proteins revealed that human b 5 residues 16 -41 were important for the specific regulation of the lyase activity of HuCYP17, possibly serving as an interacting domain with the enzyme. CYP17 may therefore have evolved from a general producer of sex steroids in lower vertebrates to a more tightly regulated producer of both sex steroids and glucocorticoids in mammals.The development and maintenance of male and female fertility depends on the normal function of androgens and their cognate androgen receptor. Androgens induce male sexual differentiation before birth, promote sexual maturation during puberty, and maintain male reproductive function in adults (1). In women, androgens serve as precursors for estrogen biosynthesis (2) while also playing significant roles in normal as well as pathologic ovarian development (3-8).Androgen production depends on the enzyme CYP17 (9). CYP17 mediates two enzymatic reactions. Its 17-hydroxylase activity primarily converts pregnenolone (⌬5 pathway) and progesterone (⌬4 pathway) to 17-hydroxypregnenolone and 17-hydroxyprogesterone, respectively. The 17,20-lyase activity of CYP17 then converts the 17-hydroxylated pregnenolone and progesterone to dehydroepiandrosterone (DHEA) 1 and androstenedione, respectively. Notably, the cofactor cytochrome b 5 has been implicated as a critical component of the 17,20-lyase reaction. Nearly all mammalian CYP17 isoforms that have been examined have little 17,20-lyase activity in the absence of b 5 . The addition of b 5 markedly enhances lyase activity, although it usually remains lower than the hydroxylase activity und...

show abstract

“…For example, corticosterone is the major glucocorticoid produced in rats and mice, whereas cortisol is the major glucocorticoid in the human, monkey, cow, rabbit, hamster and the guinea pig. This is due to the fact that guinea pigs, like humans, but unlike mouse and rat, express P450c17 and make C 19 steroids within their adrenals [19]. Due to these characteristics, the guinea pig becomes an appropriate in vivo model to study the mechanisms involved in adrenal steroidogenesis.…”

Section: Discussionmentioning

confidence: 98%

“…While corticosterone is the glucocorticoid produced in rats and mice, due to the lack of adrenal P450c17 expression, cortisol represents the major glucocorticoid in guinea pig, as in humans [19]. Although DHEA and dehydroepiandrosterone sulfate (DHEAS), being the major circulating C 19 steroids in humans, are not detected in the guinea pig circulation, it has been demonstrated that another C 19 steroid, namely 11beta-hydroxyandrostenedione (11␤-OHDIONE), is produced by the guinea pig adrenals and that the levels of circulating 11␤-OHDIONE could be stimulated after administration of adrenocorticotropic hormone (ACTH) [20][21][22][23].…”

Section: Introductionmentioning

confidence: 99%

Characterization of the guinea pig 3β-hydroxysteroid dehydrogenase/Δ5–Δ4-isomerase expressed in the adrenal gland and gonads

Durocher

Sánchez

Ricketts

et al. 2005

The Journal of Steroid Biochemistry and Molecular Biology

View full text Add to dashboard Cite

Studies of the guinea pig adrenal cytochrome P450c17 cDNA

Cited by 9 publications

References 16 publications

Comparison of the 17α-Hydroxylase/C17,20-Lyase Activities of Porcine, Guinea Pig and Bovine P450c17 Using Purified Recombinant Fusion Proteins Containing P450c17 Linked to NADPH-P450 Reductase

Comparison of the 17α-Hydroxylase/C17,20-Lyase Activities of Porcine, Guinea Pig and Bovine P450c17 Using Purified Recombinant Fusion Proteins Containing P450c17 Linked to NADPH-P450 Reductase

Xenopus laevis CYP17 Regulates Androgen Biosynthesis Independent of the Cofactor Cytochrome b5

Characterization of the guinea pig 3β-hydroxysteroid dehydrogenase/Δ5–Δ4-isomerase expressed in the adrenal gland and gonads

Contact Info

Product

Resources

About