Synthesis of alkane hydroxylase of Pseudomonas oleovorans increases the iron requirement of alk+ bacterial strains.
The alk genes enable Pseudomonas oleovorans to utilize alkanes as sole carbon and energy source. Expression of the alk genes in P. oleovorans and in two Escherichia coli recombinants induced iron limitation in minimal medium cultures. Further investigation showed that the expression of the alkB gene, encoding the integral cytoplasmic membrane protein AlkB, was responsible for the increase of the iron requirement of E. coli W3110 (pGEc47). AlkB is the non-heme iron monooxygenase component of the alkane hydroxylase system, and can be synthesized to levels up to 10% (w/w) of total cell protein in E. coli W3110 (pGEc47). Its synthesis is, however, strictly dependent on the presence of sufficient iron in the medium. Our results show that a glucose-grown E. coli alk+ strain can reach alkane hydroxylase activities of about 25 U/g cdw, and are consistent with the recent finding that catalytically active AlkB contains two, rather than one iron atom per polypeptide chain.
Staijen
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Institut für Biotechnologie, Swiss Federal Institute of Technology (ETH), ETH Hoünggerberg, HPT, 8093 Zürich, Switzerland.
Witholt
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B
D001709:Biotechnology; D003470:Culture Media; D042926:Cytochrome P-450 CYP4A; D003577:Cytochrome P-450 Enzyme System / Q000096:biosynthesis* / Q000235:genetics; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism; D005798:Genes, Bacterial; D007501:Iron / Q000378:metabolism*; D006899:Mixed Function Oxygenases / Q000096:biosynthesis* / Q000235:genetics; D011549:Pseudomonas / Q000201:enzymology* / Q000235:genetics / Q000378:metabolism; D011995:Recombination, Genetic
D003470:Culture Media; D003577:Cytochrome P-450 Enzyme System; D007501:Iron; D006899:Mixed Function Oxygenases; D042926:Cytochrome P-450 CYP4A