Sumitomo Chemical Company, Limited

Director-general of Agency of Industrial Science and Technology

Yoshiyasu Yabusaki

Hideo Ohkawa

Toshiyuki Sakaki

Koji Hayashi

Megumi Shibata

Yasushi Matsuki

Hiroko Murakami

Toshiyuki Sakai

Iwao Nakatsuka

Hideo Kaneko

Koichiro Komai

Megumi Akiyoshi

There is disclosed a method for evaluation of the safety of a chemical compound, which includes the steps of: (a) reacting a chemical compound with recombinant yeast cells expressing, or in other words producing, human cytochrome P450 molecular species P450 1A2, P450 2C9, P450 2E1 and P450 3A4 together with a yeast NADPH-P450 reductase, which may be in the form of a fused enzyme with each of said human cytochrome P450 molecular species, or with the cell free extracts of the yeast cells; and (b) analyzing the resulting metabolite to determine the safety of the compound. According to this method, it can be determined whether a test compound will be converted into a carcinogenic or mutagenic form through the metabolism in the human liver, and whether the test compound or its metabolite has mutagenicity.

Method for safety evaluation of chemical compound using recombinant yeast expressing human cytochrome P450

A flavin-containing monooxygenase comprising the amino acid sequence of SEQ ID NO:8, a microorganism producing said flavin-containing monooxygenase, a flavin-containing monooxygenase gene containing a DNA sequence coding for the amino acid sequence of SEQ ID NO:8, a flavin-containing monooxygenase gene containing the DNA sequence of SEQ ID NO:7, a plasmid containing said flavin-containing monooxygenase gene, a microorganism containing said plasmid, and a method for producing the enzyme of the present invention wherein said microorganism is cultured to produce the enzyme.

Human flavin-containing monooxygenase

The expression of mitochondrial P450 in yeast is disclosed. The mitochondrial P450 includes a chimeric P450 in which a signal sequence present at the N-terminus of a mammalian mitochondrial P450 has been substituted by a signal of a yeast mitochondrial protein and a chimeric P450 in which the latter signal has been further substituted by a targeting signal sequence to microsome. An expression plasmid for producing the enzyme in a large scale, a recombinant yeast strain carrying the expression plasmid, a process for producing the enzyme by the use of the recombinant yeast, and a process for producing 5.beta.-cholestane-3.alpha., 7.alpha., 12.alpha., 27-tetrol, 25-hydroxyvitamin D.sub.3 and 1.alpha.,25-dihydroxyvitamin D.sub.3 are also disclosed.

Mitochondrial P450

The present invention provides a chimeric enzyme gene which codes for a monooxygenase having both monooxygenase activity derived from cytochrome P-450 and reducing power supplying ability derived from NADPH-cytochrome P-450 reductase. The present invention further provides a yeast expression plasmid which contains said chimeric enzyme gene and expresses said monooxygenase gene; a transformed yeast strain which carries said yeast expression plasmid; a monooxygenase which has both the monooxygenase activity and the reducing power supplying ability as mentioned above; and a process for producing said monooxygenase.

Chimeric fused monooxygenase of cytochrome P-450 and NADPH-cytochrome

Transformed yeasts with yeast expression plasmid containing yeast alcohol dehydrogenase promoter and bovine adrenocortical cytochrome P-450.sub.17.alpha. cDNA, which produce bovine adrenocortical cytochrome P-450.sub.17.alpha., and processes for producing 17-hydroxypregnenolone and 17-hydroxyprogesterone which comprises 17-hydroxylating pregnenolone or progesterone to 17-hydroxypregnenolone or 17-hydroxyprogesterone, respectively, with the said transformed yeasts.

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Hyogo, Japan

Yoshiyasu Yabusaki