Purification and Biochemical Characterization of Taxadiene Synthase from Bacillus koreensis and Stenotrophomonas maltophilia

Taxadiene synthase (TDS) is the rate-limiting enzyme of Taxol biosynthesis that cyclizes geranylgeranyl pyrophosphate into taxadiene. Attenuating productivity by fungi main challenge impeding its industrial application; it possible silencing expression TDS most noticeable genomic feature associated with Taxol-biosynthetic abolishing in fungi. As such, characterization unique biochemical properties and autonomous independent transcriptional factors from host challenge. Thus, objective this study was to kinetically characterize endophytic bacteria isolated different plants harboring Taxol-producing Among recovered 23 isolates, Bacillus koreensis Stenotrophomonas maltophilia achieved highest activity. Upon using Plackett–Burman design, B. (18.1 µmol/mg/min) S. (14.6 increased ~2.2-fold over control. The purified gel-filtration ion-exchange chromatography ~15 overall folds molecular subunit structure 65 80 kDa maltophilia, respectively. chemical identity taxadiene authenticated GC-MS analyses, which provided same mass fragmentation pattern authentic tds gene screened PCR nested primers conservative active site domains, amplicons were sequenced, displaying a higher similarity T. baccata brevifolia. activity both bacterial isolates recorded at 37–40 °C. Apo-TDSs retained ~50% initial holoenzyme activities, ensuring their metalloproteinic identity. completely restored upon addition Mg2+, confirming Mg2+ as cofactor. dramatically reduced DTNB MBTH, implementation cysteine-reactive thiols ammonia groups on domains. This first report exploring robust affinity cyclize GGPP taxadiene, could be novel platform for production intermediary metabolites biosynthesis.