Biosynthesis and conversion of phenolics and taste active compounds in food fermentations

微生物代谢决定并维持发酵食品的质量和安全。乳酸杆菌是大多数发酵食品（包括发酵谷物食品）中发酵微生物群的主要代表。申请人的研究计划总体目标是将食品发酵乳酸杆菌的生态学和生理学与其在谷物发酵中的竞争力及其对食品质量的影响联系起来。拟议研究计划的长期目标是表征次要代谢物的代谢，特别是味觉活性肽的形成和乳酸杆菌对酚类化合物的转化，并确定这些代谢物对面包感官、营养和质地质量的影响。具体来说，该项目旨在阐明β-谷氨酰肽形成和酚酸转化的遗传决定因素，了解这些代谢途径在乳酸菌生态中的作用，及其对提高面包质量的贡献。 味觉活性 γ-谷氨酰肽通过改变甜味、咸味和鲜味化合物的感知来影响食物的味道。这种味觉活动被称为“浓味”活动。 β-谷氨酰肽在酱油、熟干酪和酸面团的发酵中积累至活性浓度；然而，它们的形成通常归因于原材料中的酶而不是细菌代谢。申请人最近的工作证明罗伊氏乳杆菌在酸面团中产生γ-谷氨酰肽。生物信息学分析鉴定出γ-谷氨酸-半胱氨酸连接酶和谷氨酰胺酶是合成γ-谷氨酰肽的酶。  α-谷氨酰肽对面包味道的贡献将通过对酸面团和酸面团面包中这些肽的定量，以及对用野生型菌株和谷氨酰胺酶或γ-谷氨酸-半胱氨酸合酶缺失的突变菌株生产的面包进行感官分析来确定。感官分析将特别旨在利用能够积累γ-谷氨酰肽的罗伊氏乳杆菌菌株来生产低盐面包而不降低消费者的偏好。谷物、蔬菜或水果中的酚酸通过其抗菌活性调节发酵微生物群的组成；此外，酚酸也是风味挥发物的前体化合物。乳酸菌的酚酸代谢对谷物发酵的生态适应性以及发酵谷物食品的质地和感官品质的贡献尚不清楚。拟议项目的工作旨在通过比较基因组学来识别对酚酸具有活性的推定酶。酚酸脱羧酶和还原酶将在异源表达后进行表征，并通过等基因缺失突变体的表征，并确定它们对酸面团生物化学和面包质量的作用。

Microbial metabolism determines and maintains the quality and safety of fermented foods. Lactobacilli are dominant representatives of fermentation microbiota in most fermented foods, including fermented cereal food. The applicant's research program generally aims to link the ecology and physiology of food-fermenting lactobacilli to their competitiveness in cereal fermentations and their impact on food quality. The long term aim of the proposed research programme is to characterize the metabolism of minor metabolites, particularly the formation of taste-active peptides and the conversion of phenolic compounds by lactobacilli, and to determine the impact of these metabolites on the sensory, nutritional, and textural quality of bread. Specifically, the proposed project aims to elucidate the genetic determinants of formation of ?-glutamyl peptides and conversion of phenolic acids, to understand the role of these metabolic pathways in the ecology of lactobacilli, and their contribution to improved bread quality. Taste active ?-glutamyl peptides impact the taste of foods by altering the perception of sweet, salty, and umami tasting compounds. This taste activity has been referred to as "kokumi" activity. ?-Glutamyl peptides accumulate to active concentrations in fermentation of soy sauce, long-ripened cheese, and in sourdough; however, their formation has typically been attributed to enzymes from the raw material rather than bacterial metabolism. Recent work of the applicant demonstrated that L. reuteri produces ?-glutamyl peptides in sourdough. Bioinformatic analyses identified ?-glutamate-cysteine ligases and glutaminases as enzymes that synthesise ?-glutamyl peptides.  The contribution of ?-glutamyl peptides to the taste of bread will be determined by quantification of these peptides in sourdough and sourdough bread, and by sensory analysis of bread produced with wild type strains and mutant strains with deletions in glutaminases or ?-glutamate-cysteine synthase. Sensory analyses will particularly aim to employ strains of L. reuteri that accumulate ?-glutamyl peptides to produce salt-reduced bread without reduction of consumer preference. Phenolic acids that in cereals, vegetables, or fruits modulate the composition of fermentation microbiota through their antimicrobial activity; moreover, phenolic acids also are precursor compounds for flavour volatiles. The contribution of phenolic acid metabolism of lactobacilli to ecological fitness in cereal fermentations and the texture and sensory quality of fermented cereal foods is unknown. Work in the proposed project aims to identify putative enzymes with activity on phenolic acids by comparative genomics. Phenolic acid decarboxylases and reductases will be characterized after heterologous expression, and by characterization of isogenic deletion mutants, and their role for sourdough biochemistry and bread quality will be determined.