Regio- and enantioselective O-demethylation of tetrahydroprotoberberines by cytochrome P450 enzyme system from Streptomyces griseus ATCC 13273.

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.

Effects of Electroacupuncture on PGC-1 α Expression in Brown Adipose Tissue.

The inducible coactivator PGC-1 α plays master regulator in mitochondrial biogenesis and thermogenesis in brown adipose tissues (BATs). BAT is a natural antiobesity organ which dissipates chemical energy in the form of heat through specialized mitochondrial protein UCP-1. Eletroacupuncture (EA) has been widely used as an alternative treatment for obesity and its related disorders such as type 2 diabetes. The molecular mechanism of electroacupuncture on treatment of obesity is still unclear. We hypothesized that electroacupuncture induced PGC-1 α expression to increase the energy expenditure in BAT. Rats were randomly divided into control group and electroacupuncture treatment group. We investigated the effects of electroacupuncture at Zusanli (ST36) acupoint on the expressions of PGC-1 α and its associated genes in the BAT of rats using real-time PCR and western blotting. We found that electroacupuncture effectively induces the expression of PGC-1 α and UCP-1 by 4-fold and 5-fold in the BAT of rats, respectively. Our results indicated that the molecular mechanism of electroacupuncture for the treatment of obesity may be, or at least partially, through induction of both PGC-1 α and UCP-1 expressions to increase energy expenditure in BAT.