Abstract
Ferulic acid decarboxylase from Saccharomyces cerevisiae (ScFDC1) was described to possess a novel, prenylated flavin mononucleotide cofactor (prFMN) providing the first enzymatic 1,3-dipolar cycloaddition mechanism. The high tolerance of the enzyme towards several non-natural substrates, combined with its high quality, atomic resolution structure nominates FDC1 an ideal candidate as flexible biocatalyst for decarboxylation reactions leading to synthetically valuable styrenes. Herein the substrate scope of ScFDC1 is explored on substituted cinnamic acids bearing different functional groups (–OCH3, –CF3 or –Br) at all positions of the phenyl ring (o−, m−, p−), as well as on several biaryl and heteroaryl cinnamic acid analogues or derivatives with extended alkyl chain. It was found that E. coli whole cells expressing recombinant ScFDC1 could transform a large variety of substrates with high conversion, including several bulky aryl and heteroaryl cinnamic acid analogues, that characterize ScFDC1 as versatile and highly efficient biocatalyst. Computational studies revealed energetically favoured inactive binding positions and limited active site accessibility for bulky and non-linear substrates, such as 2-phenylthiazol-4-yl-, phenothiazine-2-yl- and 5-(4-bromophenyl)furan-2-yl) acrylic acids. In accordance with the computational predictions, site-directed mutagenesis of residue I330 provided variants with catalytic activity towards phenothiazine-2-yl acrylic acid and provides a basis for altering the substrate specificity of ScFDC1 by structure based rational design.
Highlights
Styrenes are valuable building blocks for the synthesis of fine chemicals, polymers and pharmaceutically active compounds
Since earlier studies focused mostly on cinnamic acid derivatives with functional groups at the 4-position of the phenyl group[21], we investigated whether ScFDC1 accepts differently (o,m,p-) substituted phenyl, bulky heteroaryl- or biaryl-analogues of cinnamic acid
The tested substrate library was obtained through Knoevenagel–Doebner reaction and included (i) cinnamic acid analogues with functional groups (–Br, –OCH3 and –CF3) in different positions (o−, m−, p−) of the aromatic ring (1a–j), or (ii) substrate analogues with extended alkenyl or alkyl chains as well as (iii) several biaryl and heteroaryl analogues of cinnamic acid (1m–x) (Fig. 1)
Summary
Styrenes are valuable building blocks for the synthesis of fine chemicals, polymers and pharmaceutically active compounds Biotechnologies for their synthesis continuously emerged, styrene production from glucose through engineered Escherichia coli cells, or Saccharomyces cerevisiae cells with improved phenotypes have been successfully developed[1,2]. These methodologies rely on the activity of ferulic acid decarboxylase (FDC1) on cinnamic acid, a metabolic intermediate of the shikimate pathway. Phenylacrylic acid decarboxylases (PADs) are flavoproteins with a non-covalently bound flavin mononucleotide Their most known representative is PAD1 from E. coli, known as UbiX, which catalyse the decarboxylation of 3-octaprenyl-4-hydroxybenzoate in the ubiquinone biosynthesis[18]. Since earlier studies focused mostly on cinnamic acid derivatives with functional groups at the 4-position of the phenyl group[21], we investigated whether ScFDC1 accepts differently (o-,m-,p-) substituted phenyl-, bulky heteroaryl- or biaryl-analogues of cinnamic acid
Sign-in/Register to view highlights & summary Sign-in/Register to access full text options 
Free) 
Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a call
