Phenolic Acid Metabolism in Lactic Acid Bacteria and its Ecological Relevance
Creators: Gaur, Gautam
Publisher: name: University of Alberta Library
Year: 2022

<p>Phenolic compounds are common and structurally diverse plant secondary metabolites. Epidemiological studies have associated their consumption to health benefits such as reduction of chronic inflammation and risk of colorectal cancer. Phenolic acids are a major class of dietary phenolic compounds possessing antioxidant, antimicrobial, and bioactive properties. The metabolism of phenolic compounds by food-fermenting lactic acid bacteria is a significant contributor to the safety, nutritional content, and quality of many plant-based fermented foods. Lactobacillaceae possess enzymes responsible for hydrolysis, decarboxylation, and reduction of phenolic acids; however, several genetic determinants responsible for these activities remain uncharacterized.<br>
In this work, screening of the strain-specific phenolic acid metabolism of a diverse set of Lactobacillaceae guided a comparative genomic analysis, and enabled identification of 3 novel phenolic acid reductases and esterases. The function of par1 and hcrF from Furfurilactobacillus milii and Limosilactobacillus fermentum in reduction of hydroxycinnamic acids was confirmed via construction of deletion mutants and biochemical characterization of heterologously expressed enzyme respectively. Hydroxycinnamic acid esterase hceP was identified in Lactiplantibacillus plantarum TMW1.460; it hydrolyses chlorogenic acid and methyl ferulate. Model food fermentations with isogenic mutants of Lp. plantarum lacking phenolic acid esterases provided evidence for differential regulation of enzymes based on substrate composition, highlighting the need for confirmation of enzymatic activity in food systems for usage of strains in targeted metabolite production.<br>
To assess the role of genetic determinants in production of various phenolic acid metabolites during food fermentations, 5 isogenic mutants of Ff. milii FUA3583 lacking genes involved in hydroxycinnamic acid metabolism were created: Δest (esterase), Δpad (decarboxylase), ΔΔpar1/par2 (reductases), ΔΔΔpar1/par2/pad and ΔΔΔΔpar1/par2/pad/est. Wild type and mutant strains were used to ferment different sorghum cultivars with differences in strain behaviour and metabolite production observed upon quantification of free phenolic acid metabolites using HPLC. Competition experiments performed between mutant and wild type strains in sorghum sourdoughs indicated the contribution of phenolic acid metabolism genes to ecological fitness in a phenolic acid rich environment. Bioinformatic analysis revealed insights into the association of lifestyle and ecology to the presence of phenolic acid metabolism genes in Lactobacillaceae, with nomadic strains possessing an extensive genetic toolkit compared to almost complete lack of phenolic metabolism genes in insect-adapted lifestyles.<br>
This study also identified enzymes with unconfirmed activity homologous to characterized phenolic acid enzymes, along with documentation of metabolic activity of Lactobacillaceae on phenolic acids and patulin without known genetic determinants. These findings indicate the potential of Lactobacillaceae in metabolizing other plant and fungal secondary metabolites. Taken together, this research expands our knowledge on the behaviour and phenolic acid metabolism of Lactobacillaceae in food fermentations. The information on distribution of phenolic metabolism genes across Lactobacillaceae also allows for easier selection of strains to better control fermentation outcomes, while also documenting the potential of cereal fermentations for production of bioactive phenolics and functional foods by selectively metabolizing phenolic acids to different end products.</p>