Chemistry: molecular biology and microbiology – Enzyme – proenzyme; compositions thereof; process for... – Oxidoreductase
Reexamination Certificate
2006-04-25
2006-04-25
Patterson, Jr., Charles L. (Department: 1652)
Chemistry: molecular biology and microbiology
Enzyme , proenzyme; compositions thereof; process for...
Oxidoreductase
C435S252300, C435S320100, C536S023200
Reexamination Certificate
active
07033807
ABSTRACT:
The present invention relates to a novel cytochrome P450-like enzyme (Aspergillus ochraceus11 alpha hydroxylase) and an oxidoreductase (Aspergillus ochraceusoxidoreductase) isolated from cDNA library generated from the mRNA ofAspergillus ochraceusspores. When the cDNA encoding the 11 alpha hydroxylase was co-expressed inSpodoptera frugiperda(Sf-9) insect cells with the cDNA encoding human oxidoreductase as an electron donor, it successfully catalyzed the conversion of the steroid substrate 4-androstene-3,17-dione (AD) to 11 alpha-hydroxy-AD as determined by HPLC analysis. The invention also relates to nucleic acid molecules associated with or derived from these cDNAs including complements, homologues and fragments thereof, and methods of using these nucleic acid molecules, to generate, for example, polypeptides and fragments thereof. The invention also relates to the generation of antibodies that recognizes theA. ochraceus11 alpha hydroxylase and oxidoreductase and methods of using these antibodies to detect the presence of these native and recombinant polypeptides within unmodified and transformed host cells, respectively. The invention also provides methods of expressing theAspergillus11 alpha hydroxylase gene separately, or in combination with human orAspergillusoxidoreductase, in heterologous host cells, to facilitate the bioconversion of steroid substrates to their 11 alpha hydroxy-counterparts.
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Bolten Suzanne L.
Clayton Robert A.
Easton Alan M.
Engel Leslie C.
Messing Dean M.
Patterson Jr. Charles L.
Pharmacia Corporation
Slavinsky Christopher W.
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