Functional Properties of Peptides Produced by Bacterial Enzymes from Traditional Fermented Foods

几千年来，人们通过传统和自然发酵来制备食物。天然存在的微生物如细菌、酵母和霉菌产生蛋白水解酶用于制造食物。多种芽孢杆菌参与了传统的发酵谷物和豆类食品。本研究旨在从发酵食品中收集微生物，并表征细菌酶的蛋白水解潜力和功能特性。根据《生物多样性公约》，与泰国、越南、中国和柬埔寨签署了合作实验室谅解备忘录，因此，这项研究的数据可以公开分发。对分离到的细菌进行16S rDNA鉴定，并进行遗传树分析。纯化酶的分子量为33 kDa，N端氨基酸序列为AQSVPYGISQIKAPA。然而，这是第一次从传统发酵小麦面食品中分离到它。纯化后的蛋白酶能将酸性酪蛋白酶解成C-末端含有亲水性和疏水性氨基酸的片段，特别是Arg、Glu、Val和Ile。枯草杆菌DB和SR可通过水解小麦过敏原麦醇溶蛋白片段来降低麦醇溶蛋白的致敏性。此外，B.枯草芽孢杆菌DB也能在1h内将β-乳球蛋白（牛奶过敏原）消化成小片段，这些片段在孵育3h后消失。然后分析片段的N-末端氨基酸序列，发现该酶可水解β-乳球蛋白的主要过敏原表位Gln ^-Ser ^。<35><36>因此，这些酶可用于生产低过敏性小麦粉或奶食品。

For many thousands of years, people have prepared foods by traditional and natural fermentation. Naturally occurring microorganisms such as bacteria, yeast and moulds produce the proteolytic enzymes for making foods. A variety of Bacillus sp. have been involved in traditional fermented cereal and legume foods. This study aimed to collect microorganisms from fermented foods and characterize proteolytic potentials and functional properties of bacterial enzymes. Memorandum of Understanding (MOU) of collaboratory based on the Convention on Biological Diversity was decided with Thailand, Vietnam, China and Cambodia, the data from this research could therefore be publicly distributed. The isolated bacteria were identified by 16S rDNA and phylogenetic-tree analysis was also performed. The molecular mass and N-terminal amino acid sequence of purified enzyme were 33 kDa and AQSVPYGISQIKAPA, respectively. However, this was the first report in which it was isolated from traditional fermented wheat flour food. The purified protease digested acid casein into fragments with hydrophilic and hydrophobic amino acids at C-terminal, in particular Arg, Glu, Val and Ile.The concentrated crude enzyme of B. subtilis DB and SR could reduce allergenicity of gliadin (wheat allergen) by hydrolyzing the allergenic gliadin fragments detected by immunoblotting. Furthermore, the concentrated crude enzyme of B. subtilis DB could also digest β-lactoglobulin (milk allergen) into small fragments within 1 h and these fragments were disappeared after 3-h incubation. The fragments were then analyzed for N-terminal amino acid sequence ; it was found that this enzyme could hydrolyze the major allergenic epitope of β-lactoglobulin at Gln^<35>-Ser^<36> position. Therefore these enzymes can be applied for the production of hypoallergenic wheat flour or milk food products.