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Amino acid substitutions Phe66----Leu and Ser126----Pro abolish cortisol and aldosterone synthesis by bovine cytochrome P450(11)beta.
Mathew PA, Mason JI, Trant JM, Sanders D, Waterman MR
(1990) J Biol Chem 265: 20228-33
MeSH Terms: Adrenal Cortex, Aldosterone, Animals, Base Sequence, Cattle, Cell Line, Cloning, Molecular, Codon, DNA, Desoxycorticosterone, Hydrocortisone, Leucine, Mitochondria, Molecular Sequence Data, Mutagenesis, Site-Directed, Oligonucleotide Probes, Phenylalanine, Plasmids, Polymerase Chain Reaction, Proline, RNA, Messenger, Serine, Steroid 11-beta-Hydroxylase, Transfection
Show Abstract · Added February 12, 2015
A cDNA clone encoding the complete protein sequence of the precursor form of bovine cytochrome P450(11)beta has been constructed using a combined technique of first strand cDNA synthesis by reverse transcription followed by polymerase chain reaction. Upon expression of this cDNA in COS 1 cells the P450(11)beta is found to be proteolytically processed and localized in the mitochondrion. This cDNA encodes the major form of P450(11)beta found in bovine adrenal cortex (designated 11 beta-3; Kirita, S., Morohashi, K., Hashimoto, T., Yoshioka, H., Fujii-Kuriyama, Y., and Omura, T. (1988) J. Biochem. 104, 683-686) and is capable of catalyzing 11 beta-hydroxylation of deoxycorticosterone, 11-deoxycortisol, and androstenedione in COS 1 cells as well as aldosterone synthesis from deoxycorticosterone. In addition, a second form of P450(11)beta (herein designated 11 beta-4), having no detectable 11 beta-hydroxylase activity or aldosterone synthase activity, was found in the local bovine population by this cloning procedure. These two forms of P450(11)beta (11 beta-3 and 11 beta-4) contain five amino acid differences between them, all located within the amino-terminal half of the molecules. By changing 2 of the amino acids in the inactive form to the corresponding amino acids in the active form (Leu66----Phe and Pro126----Ser) both 11 beta-hydroxylase and aldosterone synthetase activities were completely restored. Neither of these changes alone led to detectable activity. Thus, upon expression in mitochondria of heterologous cells, bovine P450(11)beta catalyzes both 11 beta-hydroxylation and aldosterone synthesis as reported previously for the purified enzyme in an in vitro reconstituted system, and Phe66 and Ser126 seem to be important residues in maintaining both activities.
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24 MeSH Terms
Regulation of the prefrontal cortical dopamine system by the neuroactive steroid 3a,21-dihydroxy-5a-pregnane-20-one.
Grobin AC, Roth RH, Deutch AY
(1992) Brain Res 578: 351-6
MeSH Terms: 3,4-Dihydroxyphenylacetic Acid, Anesthetics, Animals, Cerebral Cortex, Cerebral Ventricles, Desoxycorticosterone, Dopamine, Dose-Response Relationship, Drug, Exploratory Behavior, Homovanillic Acid, Injections, Intraventricular, Male, Motor Activity, Rats, Rats, Inbred Strains, Reference Values, Stress, Psychological
Show Abstract · Added May 27, 2014
The dopamine innervation of the prefrontal cortex can be differentiated from other telencephalic dopamine projection fields by its sensitivity to stress. The stress-induced activation of the mesoprefrontal cortical dopamine system can be blocked by pretreatment with benzodiazepines. A group of neuroactive steroids that modulate GABA-induced chloride flux through means distinct from that of the benzodiazepines has recently been identified. Intraventricular administration of the neuroactive steroid 3 alpha,21-dihydroxy-5 alpha-pregnane-20-one resulted in a dose-dependent decrease in dopamine metabolites in the prefrontal cortex, but not in mesolimbic or striatal sites; sedative effects were not observed. Moreover, the neuroactive steroid selectively attenuated the stress-induced activation of the mesoprefrontal cortical dopamine system. These data suggest that neuroactive steroids may function as endogenous anxiolytic agents.
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17 MeSH Terms