Identification of a multi-component berberine 11-hydroxylase from Burkholderia sp. strain CJ1.

Berberine (BBR) is a protoberberine alkaloid extracted from plants such as Coptis japonica (Ranunculaceae). In a previous report, we demonstrated the existence of a 11-hydroxylation pathway employed by BBR-utilizing bacteria for metabolism of BBR. In the present study, we report the identification of the genes brhA, brhB, and brhC as encoding a multicomponent BBR 11-hydroxylase in Burkholderia sp. strain CJ1. BrhA is belonging to the Rieske non-heme iron oxygenase (RO) family, a class of enzymes known to catalyze the first step in bacterial aromatic-ring hydroxylation. We further demonstrate that BrhA activity requires BrhB (ferredoxin reductase) and BrhC (ferredoxin) as electron transport chain components. A BLAST search revealed that BrhA exhibits 38% and 33% sequence identity to dicamba O-demethylase (DdmC; AY786443) and chloroacetanilide herbicides N-dealkylase (CndA; KJ461679), respectively. To our knowledge, this work represents the first report of a bacterial oxygenase catalyzing the metabolism of a polycyclic aromatic-ring alkaloid.Abbreviations: BBR: berberine; D-BBR: demethyleneberberine; H-BBR: 11-hydroxyberberine; HD-BBR: 11-hydroxydemethyleneberberine; HDBA: 2-hydroxy-3,4-dimethoxybenzeneacetic acid; PAL: palmatine; H-PAL: 11-hydroxypalmatine; BRU: berberrubine; Fd: ferredoxin; FdR: ferredoxin reductase; ETC: electron transport chain.

Characterization of Burkholderia sp. strain CJ1, a newly isolated berberine-degrading bacterium from rhizosphere of Coptis japonica.

Burkholderia sp. strain CJ1 was newly isolated as berberine (BBR) degrading bacteria from rhizosphere of Coptis japonica. CJ1 had the ability to utilize BBR as the sole carbon source and revealed that BBR metabolism via 11-hydroxylation and demethylenation pathway. It was also revealed that the 11-hydroxylation ability of BBR and palmatine (PAL) has induced by BBR.

Antimicrobial agent isolated from Coptidis rhizome extract incubated with Rhodococcus sp. strain BD7100.

Coptidis rhizome (CR) is a widely used herbal medicine that contains protoberberine-type alkaloids. CR extract exhibits various pharmacologic activities. A previous study reported the isolation of Rhodococcus sp. strain BD7100 as a berberine (BBR)-utilizing bacterium, and the BBR-degradation pathway has been investigated. When we incubated strain BD7100 cells with CR extract, the number of viable cells declined with the degradation of components in the CR extract, and the culture broth exhibited antibacterial activity against strain BD7100. These results suggest that CR extract cultured in the presence of strain BD7100 contains one or more antibacterial agents. In this study, we isolated coptirhoquinone A (1) from CR extract incubated with strain BD7100 in Luria-Bertani (LB) medium, and the structure was elucidated using NMR and MS analysis. We also report the total synthesis and antimicrobial activities of 1 against bacteria, fungi, and Pythium sp.

Common origin of methylenedioxy ring degradation and demethylation in bacteria.

Plants produce many specific secondary metabolites as a response to environmental stress, especially biological stress. These compounds show strong biological activities and high stability against degradation by microbes and animals. Berberine, a benzylisoquinoline alkaloid, is found in many plant species and has strong antimicrobial activity, and is often included in traditional herbal medicines. We previously investigated how berberine is degraded in nature and we isolated two berberine-utilizing bacteria. In this study, we characterized the gene encoding the enzyme that degrades the 2,3-methylenedioxy ring of berberine; this ring is important for its activity and stability. Further characterization of several other berberine-utilizing bacteria and the genes encoding key demethylenation enzymes revealed that these enzymes are tetrahydrofolate dependent and similar to demethylation enzymes such as GcvT. Because the degradation of O-methyl groups or the methylenedioxy ring in phenolic compounds such as lignin, lignan and many other natural products, including berberine, is the key step for the catabolism of these compounds, our discovery reveals the common origin of the catabolism of these stable chemicals in bacteria.