Generation of 4-vinylguaiacol through a novel high-affinity ferulic acid decarboxylase to obtain smoke flavours without carcinogenic contaminants.

Thorben Detering,Katharina Mundry,Ralf G Berger

doi:10.1371/journal.pone.0244290

Abstract

Traditional smoke flavours bear the risk of containing a multitude of contaminating carcinogenic side-products. Enzymatic decarboxylation of ferulic acid released from agro-industrial side-streams by ferulic acid esterases (FAE) enables the sustainable generation of pure, food grade 4-vinylguaiacol (4-VG), the impact compound of smoke flavour. The first basidiomycetous ferulic acid decarboxylase (FAD) was isolated from Schizophyllum commune (ScoFAD) and heterologously produced by Komagataella phaffii. It showed a molecular mass of 21 kDa, catalytic optima at pH 5.5 and 35°C, and a sequence identity of 63.6% to its next relative, a FAD from the ascomycete Cordyceps farinosa. The ScoFAD exhibited a high affinity to its only known substrate ferulic acid (FA) of 0.16 mmol L-1 and a turnover number of 750 s-1. The resulting catalytic efficiency kcat KM-1 of 4,779 L s-1 mmol-1 exceeded the next best known enzyme by more than a factor of 50. Immobilised on AminoLink Plus Agarose, ScoFAD maintained its activity for several days. The combination with FAEs and agro-industrial side-streams paves the way for a new generation of sustainable, clean, and safe smoke flavours.

Highlights

The growing customer demand for natural flavours goes hand in hand with new enzymatic tools providing an alternative access to flavour compounds, which were formerly synthesised from crude oil by often harsh methods
A novel ferulic acid decarboxylase to obtain smoke flavours were purchased from Thermo Fisher Scientific
Inducing the expression of the ScoFAD encoding gene, 25 mL of the pre-culture was added to 250 mL 100 mL L-1 SNL containing 10 g L-1 wheat bran

Summary

Introduction

The growing customer demand for natural flavours goes hand in hand with new enzymatic tools providing an alternative access to flavour compounds, which were formerly synthesised from crude oil by often harsh methods. Basidiomycetes, one of the two main divisions of higher fungi, offer a great and almost untouched diversity of biocatalysts. Growing on wood, one of the most recalcitrant natural materials, xylophilic fungi feature potent oxidoreductases and hydrolases, such as laccases and phenolic acid esterases [1, 2]. Many basidiomycetes produce various and sometimes unique aroma compounds. Besides the well-known champignon flavour impact compound 1-octen-3-ol, they are capable of producing many other and sometimes rather unusual flavours, such as the 5,7,9-decatrienones with pineapple notes and (+)-nootkatone with grapefruit notes [3, 4].

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