Abstract
Tetrahydroprotoberberines (THPBs), a class of naturally occurring isoquinoline alkaloids, contain substituent methoxyl or hydroxyl groups which play a significant role in the pharmacological properties of these molecules. In this study, we report a biocatalytic strategy for selective O-demethylation of THPBs. CYP105D1, a cytochrome P450 from Streptomyces griseus ATCC 13273, exhibited markedly regioselective demethylation of nonhydroxyl-THPBs and monohydroxyl-THPBs on the D-ring. A possible binding mode of THPBs with CYP105D1 was investigated by docking analysis, and the results revealed that the D-rings of THPBs were with the minimum distance to the heme iron. Tetrahydropalmatine was used as a model substrate and enantioselective demethylation was demonstrated. (S)-Tetrahydropalmatine was only demethylated at C-10, while (R)-tetrahydropalmatine was first demethylated at C-10 and then subsequently demethylated at C-9. The kcat/Km value for demethylation of (R)-tetrahydropalmatine by CYP105D1 was 3.7 times greater than that for demethylation of (S)-tetrahydropalmatine. Furthermore, selective demethylation of (S)-tetrahydropalmatine by the CYP105D1-based whole-cell system was demonstrated for the highly efficient production of (S)-corydalmine which has distinct pharmacological applications, such as providing relief from bone cancer pain and reducing morphine tolerance. Moreover, a homologous redox partner was identified to enhance the catalytic efficiency of the CYP105D1-based whole-cell system. This is the first enzymatic characterization of a cytochrome P450 that has regio- and enantioselective demethylation activity of THPBs for application purpose. The cytochrome P450 system could be a promising strategy for selective demethylation in the pharmaceutical industry.
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This work was supported by the Major Scientific and Technological Specialized Project for “New Drugs Development” (No. 2012ZX09J12110-06B) and the Priority Academic Program Development of Jiangsu Higher Education Institutions (PAPD).
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Shen, C., Shan, T., Zhao, W. et al. Regio- and enantioselective O-demethylation of tetrahydroprotoberberines by cytochrome P450 enzyme system from Streptomyces griseus ATCC 13273. Appl Microbiol Biotechnol 103, 761–776 (2019). https://doi.org/10.1007/s00253-018-9416-4
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DOI: https://doi.org/10.1007/s00253-018-9416-4