Vavricka, Christopher J.Yoshida, TakanobuKuriya, YukiTakahashi, ShunsukeOgawa, TeppeiOno, FumieAgari, KazukoKiyota, HiromasaLi, JianyongIshii, JunTsuge, KenjiMinami, HiromichiAraki, MichihiroHasunuma, TomohisaKondo, Akihiko2019-07-302019-07-302019-05-012041-17232015http://hdl.handle.net/10919/92047Previous studies have utilized monoamine oxidase (MAO) and L-3,4-dihydroxyphenylalanine decarboxylase (DDC) for microbe-based production of tetrahydropapaveroline (THP), a benzylisoquinoline alkaloid (BIA) precursor to opioid analgesics. In the current study, a phylogenetically distinct Bombyx mori 3,4-dihydroxyphenylacetaldehyde synthase (DHPAAS) is identified to bypass MAO and DDC for direct production of 3,4-dihydroxyphenylacetaldehyde (DHPAA) from L-3,4-dihydroxyphenylalanine (L-DOPA). Structure-based enzyme engineering of DHPAAS results in bifunctional switching between aldehyde synthase and decarboxylase activities. Output of dopamine and DHPAA products is fine-tuned by engineered DHPAAS variants with Phe79Tyr, Tyr80Phe and Asn192His catalytic substitutions. Balance of dopamine and DHPAA products enables improved THP biosynthesis via a symmetrical pathway in Escherichia coli. Rationally engineered insect DHPAAS produces (R,S)-THP in a single enzyme system directly from L-DOPA both in vitro and in vivo, at higher yields than that of the wild-type enzyme. However, DHPAAS-mediated downstream BIA production requires further improvement.enCreative Commons Attribution 4.0 InternationalArticle - Refereedhttps://doi.org/10.1038/s41467-019-09610-21031043610