Biocatalytic in Vitro and in Vivo FMN Prenylation and (De)carboxylase Activation

Khorcheska A. Batyrova, Anna N. Khusnutdinova, Po Hsiang Wang, Rosa Di Leo, Robert Flick, Elizabeth A. Edwards, Alexei Savchenko, Alexander F. Yakunin

研究成果: 雜誌貢獻期刊論文同行評審

11 引文 斯高帕斯(Scopus)


Reversible UbiD-like (de)carboxylases represent a large family of mostly uncharacterized enzymes, which require the recently discovered prenylated FMN (prFMN) cofactor for activity. Functional characterization of novel UbiDs is hampered by a lack of robust protocols for prFMN generation and UbiD activation. Here, we report two systems for in vitro and in vivo FMN prenylation and UbiD activation under aerobic conditions. The in vitro one-pot prFMN cascade includes five enzymes: FMN prenyltransferase (UbiX), prenol kinase, polyphosphate kinase, formate dehydrogenase, and FMN reductase, which use prenol, polyphosphate, formate, ATP, NAD+, and FMN as substrates and cofactors. Under aerobic conditions, this cascade produced prFMN from FMN with over 98% conversion and activated purified ferulic acid decarboxylase Fdc1 from Aspergillus niger and protocatechuic acid decarboxylase ENC0058 from Enterobacter cloaceae. The in vivo system for FMN prenylation and UbiD activation is based on the coexpression of Fdc1 and UbiX in Escherichia coli cells under aerobic conditions in the presence of prenol. The in vitro and in vivo FMN prenylation cascades will facilitate functional characterization of novel UbiDs and their applications.

頁(從 - 到)1874-1882
期刊ACS Chemical Biology
出版狀態已出版 - 17 7月 2020


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