| Abstrakt: |
A recombinant Escherichia coliexpressing P450pyrmonooxygenase of Sphingomonassp. HXN‐200 was developed as a useful biocatalyst for regio‐ and stereoselective hydroxylations, with no side reaction and easy cell growth. The resting E. colicells showed an activity of 4.1 U/g cdw and 9.9 U/g cdw for the hydroxylation of N‐benzylpyrrolidin‐2‐one 1and N‐benzyloxycarbonylpyrrolidine 3, respectively, being as active as the wide‐type strain. Biohydroxylation of N‐benzylpyrrolidin‐2‐one 1with the resting cells gave (S)‐N‐benzyl‐4‐hydroxypyrrolidin‐2‐one 2in >99% eeand 10.8 mM, a 2.6 times increase of product concentration in comparison with the wild‐type strain. Biohydroxylation of N‐tert‐butoxycarbonylpiperidin‐2‐one 5, N‐benzylpiperidine 7and N‐tert‐butoxycarbonylazetidine 9with the E. colicells afforded the corresponding 4‐hydroxypiperidin‐2‐one 6, 4‐hydroxypiperidine 8, and 3‐hydroxyazetidine 10in 14 mM, 17 mM, and 21 mM, respectively. Moreover, hydroxylation of (−)‐β‐pinene 11with the recombinant E. colicells showed excellent regio‐ and stereoselectivity and gave (1R)‐trans‐pinocarveol 12in 82% yield and 4.1 mM, which is over 200 times higher than that obtained with the best biocatalytic system known thus far. The recombinant strain was also able to hydroxylate other types of substrates with unique selectivity: biohydroxylation of norbornane 13gave exo‐norbornaeol 14, with exo/endoselectivity of 95%; tetralin 15and 6‐methoxytetralin 17were hydroxylated at the non‐activated 2‐position, for the first time, with regioselectivities of 83–84%. |
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