Abstract
BackgroundThe yeast amino acid catabolism plays an important role in flavour generation since higher alcohols and acetate esters, amino acid catabolism end products, are key components of overall flavour and aroma in fermented products. Comparative studies have shown that other Saccharomyces species, such as S. kudriavzevii, differ during the production of aroma-active higher alcohols and their esters compared to S. cerevisiae.ResultsIn this study, we performed a comparative analysis of the enzymes involved in the amino acid catabolism of S. kudriavzevii with their potential to improve the flavour production capacity of S. cerevisiae. In silico screening, based on the severity of amino acid substitutions evaluated by Grantham matrix, revealed four candidates, of which S. kudriavzevii Aro10p (SkAro10p) had the highest score. The analysis of higher alcohols and esters produced by S. cerevisiae then revealed enhanced formation of isobutanol, isoamyl alcohol and their esters when endogenous ARO10 was replaced with ARO10 from S. kudriavzevii. Also, significant differences in the aroma profile were found in fermentations of synthetic wine must. Substrate specificities of SkAro10p were compared with those of S. cerevisiae Aro10p (ScAro10p) by their expression in a 2-keto acid decarboxylase-null S. cerevisiae strain. Unlike the cell extracts with expressed ScAro10p which showed greater activity for phenylpyruvate, which suggests this phenylalanine-derivative to be the preferred substrate, the decarboxylation activities measured in the cell extracts with SkAro10p ranged with all the tested substrates at the same level. The activities of SkAro10p towards substrates (except phenylpyruvate) were higher than of those for ScAro10p.ConclusionsThe results indicate that the amino acid variations observed between the orthologues decarboxylases encoded by SkARO10 and ScARO10 could be the reason for the distinct enzyme properties, which possibly lead to the enhanced production of several flavour compounds. The knowledge on the important enzyme involved in higher alcohols biosynthesis by S. kudriavzevii could be of scientific as well as of applied interest.Electronic supplementary materialThe online version of this article (doi:10.1186/s12934-016-0449-z) contains supplementary material, which is available to authorized users.
Highlights
The yeast amino acid catabolism plays an important role in flavour generation since higher alcohols and acetate esters, amino acid catabolism end products, are key components of overall flavour and aroma in fer‐ mented products
In silico analysis revealed the largest amount of radical amino acid substitutions between the phenylpyruvate decarboxylase (Aro10p) orthologues To perform a comparative analysis, DNA sequences of the orthologue genes encoding 23 enzymes which are involved in amino acid catabolism leading to higher alcohols and acetate ester formation were obtained from 75 S. cerevisiae strains and two S. kudriavzevii strains, all available in databases (Additional File 1)
The individual changes in S. kudriavzevii sequences were quantified by Grantham matrix, which scores the difference between two amino acids according to composition, polarity and molecular volume
Summary
The yeast amino acid catabolism plays an important role in flavour generation since higher alcohols and acetate esters, amino acid catabolism end products, are key components of overall flavour and aroma in fer‐ mented products. In Saccharomyces cerevisiae, the predominant yeast in foodrelated fermentations, depending on the conditions these higher alcohols are synthesised from 2-keto acids derived either from glycolysis or from the catabolism of valine, leucine and phenylalanine [4,5,6,7] on a reaction pathway known as the Ehrlich pathway [8, 9] On this pathway, the amino acids, which are transported by amino acid permeases (codified by GAP1, BAP2, BAP3, MUP3) [10,11,12,13] are first transaminated to the corresponding 2-keto acids by transaminases (codified by BAT1, BAT2, ARO8, ARO9) [14,15,16]. Acetate ester breakdown is affected by the function of hydrolases, such as those encoded by Iah1p [21] which, together with Atf1p and Atf2p, maintain an optimal ester accumulation rate
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