1991
DOI: 10.1016/0014-5793(91)80333-x
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Expression of rat liver vitamin D3 25‐hydroxylase cDNA in Saccharomyces cerevisiae

Abstract: The cDNA coding for the precursor protein of rat liver mitochondrial vitamin D3 25‐hydroxylase, cytochrome P450LMT25, was expressed under the control of the yeast alcohol dehydrogenase I promoter and terminator in Saccharomyces cerevisiae AH22 cells. The transformed years cells produced a P450LMT25 protein with an almost similar apparent molecular weight as compared with that of the authentic mature enzyme. The expression level of the P450LMT25 hemoprotein was about 5 × 104 molecules per cell as determined by … Show more

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Cited by 37 publications
(8 citation statements)
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“…No peak was observed around 450 nm from a transformant harboring a plasmid without an AoCYP, indicating that the recombinant AoCYP enzyme was successfully expressed in S. cerevisiae in an active form. These results also implied that endogenous CYPs of S. cerevisiae did not interfere with spectroscopic analysis of recombinant AoCYPs due to their low levels of expression in this host strain (2). We have confirmed significant expression of 84 AoCYPs based upon CO difference spectra.…”
supporting
confidence: 65%
“…No peak was observed around 450 nm from a transformant harboring a plasmid without an AoCYP, indicating that the recombinant AoCYP enzyme was successfully expressed in S. cerevisiae in an active form. These results also implied that endogenous CYPs of S. cerevisiae did not interfere with spectroscopic analysis of recombinant AoCYPs due to their low levels of expression in this host strain (2). We have confirmed significant expression of 84 AoCYPs based upon CO difference spectra.…”
supporting
confidence: 65%
“…CYP27A1, also known as the sterol 27-hydroxylase, is a key enzyme in bile acid formation (6, 7). Recombinant CYP27A1 is able to catalyze multiple oxidation reactions with broad substrate specificity in vitro (8)(9)(10)(11)(12)(13)(14). However, it is a lowaffinity, high-capacity vitamin D 25-hydroxylase and is certainly involved in steroidogenesis.…”
mentioning
confidence: 99%
“…Incubation of pig liver mitochondria with isocitrate and 25-hydroxyvitamin D3 resulted in the formation of both la,25-dihydroxyvitamin D3 and 25,27-dihydroxyvitamin D3 (Table 1). CYP27 is known to catalyze the 27-hydroxylation of 25-hydroxyvitamin D3 (7) and the 25-hydroxylation of vitamin D3 (5,(9)(10)(11). In further purification of the liver mitochondrial cytochrome P450 catalyzing la-hydroxylation, these activities were monitored for comparison.…”
Section: Resultsmentioning
confidence: 99%
“…The enzymes were not purified or further characterized. The pig, rabbit, and rat liver mitochondrial sterol 27-hydroxylase (CYP27) is known to catalyze 24-, 25-, and 27-hydroxylations of C27 steroids and vitamin D3 compounds (4)(5)(6)(7)(8)(9)(10)(11)(12). cDNA encoding the human liver CYP27 was isolated by Cali and Russell (13).…”
mentioning
confidence: 99%