Heme-oxygenase-like protein, FlcE undergoes a four-electron oxidation to effect multiple turnovers in the absence of any reductant
Research Poster Physical Sciences & Mathematics 2025 Graduate ExhibitionPresentation by Sohini Saha
Exhibition Number 219
Abstract
Heme-oxygenase-like diiron oxidases and oxygenases (HDOs) are an emerging superfamily of dioxygen-activating non-heme diiron enzymes. In terms of functional diversity, initially characterized members represented just two functional subclasses: the N-oxygenases and the C-C-bond-fragmenting desaturases. More recently, a new HDO, FlcE, has been discovered in the fluopsin C biosynthetic pathway, and it has been shown to catalyze both N-oxygenation and decarboxylation of (2R)-{[(2R)-2-amino-2-carboxyethyl]sulfanyl}butanedioic acid [(2R)-S-succinyl-L-cysteine)] transforming it to (2R)-{[2-hydroxyimino-ethyl]sulfanyl}butanedioic acid. While reactions with only oxidative decarboxylation or N-oxygenation outcomes have previously been observed for HDOs, both reactions occurring together is unique, thus giving rise to speculation of a novel mechanistic route. Insights into the stoichiometry and behavior of this unique reaction was afforded with the help of quantitative LC-MS analysis. In this poster, we will also be demonstrating the spectroscopic characterization of the intermediates involved in the transformation with the help of stopped-flow absorption spectroscopy and Mössbauer spectroscopy, and correlate them with the previously published N-oxygenation mechanisms of members of the ferritin-like non-heme diiron oxygenase family. This study reports the first occurrence of a fully self-reliant multi-turnover protein in the HDO superfamily and elucidates its catalytic mechanism without the aid of an external reducing system.
Importance
Recently, enzymes of aldoxime-nitrile pathway are receiving attention for their potential role in organic synthesis because of opportunities that were unprecedented in biocatalysis and green chemistry. In addition to industrial synthesis, co-expression in plant systems to improve agronomical characteristics of the crop and horticulture are also being explored. Since chemical synthesis of groups like aldoximes, amides, nitrile and acids require extreme reaction conditions like high temperature, high pressure, and energy intensive steps along with the generation of toxic side products, it is the need of the hour to address organic synthesis with future biocatalytic routes to be ecofriendly and economical. Therefore, enzymes participating in such pathways need to be explored, studied, improved and applied for organic synthesis at large scale.