CN114585258A - 用于控制真菌腐败的细菌组合物和其用途 - Google Patents

用于控制真菌腐败的细菌组合物和其用途 Download PDF

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CN114585258A
CN114585258A CN202080073495.4A CN202080073495A CN114585258A CN 114585258 A CN114585258 A CN 114585258A CN 202080073495 A CN202080073495 A CN 202080073495A CN 114585258 A CN114585258 A CN 114585258A
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A·戈尔
C·L·M·尼尔森
赫勒·斯科沃·古尔达格
S·K·迪默
K·L·V·安德森
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Abstract

本发明涉及一种包含含锰转运蛋白的乳酸菌的直投式起子培养物组合物，其用于发酵食物产品和用于抑制或延迟所述食物产品中的真菌生长，其特征在于，所述组合物包含至多600ppm的锰。

Description

用于控制真菌腐败的细菌组合物和其用途
技术领域
本发明涉及高度浓缩的起子培养物组合物和其制剂，其用于控制真菌腐败而不借助于防腐剂，诸如化学防腐剂。因此，本发明有助于满足对较少加工且不含防腐剂的食品的需求，而同时有助于提供一种有效的解决方案，以管理酵母和霉菌生长。本发明还涉及包含所述组合物的食物产品。
背景技术
食品工业中的一个主要问题是有害微生物引起的腐败。根据粮食和农业组织(Food and Agriculture Organization,FAO)，预定供人消耗的每4卡路里中就有1卡路里最终不被人消耗。在超过8亿人遭受饥饿的食物短缺时代，食物浪费话题已成为全球政策制定者和食品制造商的优先考虑问题。除了对社会造成负面的社会和经济影响之外，浪费的食物还造成一系列相关的环境影响，包括不必要的温室气体排放以及对诸如水和土地的稀缺资源的低效利用。
酵母和霉菌在引起食物腐败方面高度有效，并且是大多数食品制造商所面临的问题。由于酵母和霉菌引起的腐败在食物产品表面上清晰可见为霉菌块或变色，允许其在食用前被处理掉。酵母倾向于以浮游形式在食物和饮料基质内生长，并且它们倾向于使糖发酵，在厌氧条件下生长良好。相比之下，霉菌倾向于以由细胞组成的可见菌丝体的形状在产品表面上生长。
抗真菌微生物剂用于食物生物保存的用途是已知的，例如描述于Salas等“用于食物生物保存的抗真菌微生物剂——综述(Antifungal microbial agents for foodbiopreservation—a review).”Microorganisms 5.3(2017):37中。
特别是在乳品行业，欧洲每年有2900万吨乳制品被浪费。保持乳制品新鲜的主要挑战之一是管理酵母和霉菌的污染，酵母和霉菌天然存在于任何地方，尤其是在从生产到消费者餐桌的冷链出现中断的情况下。
出于经济和环境原因，一直需要有效控制酵母和霉菌污染的新颖或改善的方式。
发明内容
锰已被认为对人体健康至关重要，因此是一种必需的痕量元素。锰对人和动物两者的正常功能都是必不可少的，因为它是许多细胞酶(例如锰超氧化物歧化酶、丙酮酸羧化酶)的功能所必需的，并且它可用于活化许多其他酶，如激酶、脱羧酶、转移酶和水解酶。
锰可天然存在于许多食物来源中，包括叶菜、坚果、谷物和动物产品。常见食品中锰浓度的典型范围是例如谷物产品中0.4-40ppm，肉类、家禽、鱼和蛋中0.1-4ppm，蔬菜产品中0.4-7ppm，均质化乳中0.03ppm。
除了作为膳食补充剂之外，有时还在发酵产品中添加锰作为活性成分以促进双歧杆菌在乳中的生长(参见例如WO2017/021754,Compagnie Gervais Danone,France)。
最近发现，锰是食物产品中真菌生长的重要生长限制因素。因此，有可能在食物产品中施加锰清除剂，诸如锰清除细菌，以与真菌竞争游离锰。这导致这种营养素的耗尽，从而抑制或延迟真菌生长。此类细菌可用作用于发酵食物产品的起子培养物。本发明涉及此类用于工业应用的抗真菌细菌的制备。
本发明部分地基于令人惊讶的发现，即当在按比例放大工艺期间在制备起子培养物中使用高水平的锰(这是本领域中的常见实践)时，当稍后应用于食物产品中时，细菌在抑制或延迟真菌生长方面变得不太有效。因此，按比例放大工艺期间的细菌不应暴露于高水平的锰，因为此类水平会对其抗真菌活性产生负面影响。换句话说，本发明人已发现细菌组合物中所含的锰水平与其抗真菌活性密切相关。
本发明涉及高度浓缩的生物质组合物，其包含抑制酵母或霉菌生长的单一或多种乳酸菌菌株。生物质组合物可通过在生长培养基中培育细菌并提高细菌浓度来制备。
商业起子培养物通常可以冷冻或冷冻干燥(FD)培养物的形式分配。高度浓缩的培养物在商业上非常引人关注，因为此类培养物可直接接种到乳中而无需中间转移。高度浓缩的培养物可被称为直投式(direct vat set,DVS)-培养物。
商业上高度浓缩的DVS起子培养物可作为呈粉末形式的冷冻干燥或冻干的培养物。在这种形式中，起子可在没有冷藏的情况下运输。
乳酸菌通常以用于批量起子繁殖的冷冻或冷冻干燥培养物的形式供应给食品工业，诸如乳品业，或以预定用于直接接种到发酵容器或桶中的所谓的“直投式”(DVS)培养物的形式供应用于生产乳制品，诸如发酵乳产品或奶酪，而无需制备批量起子。
直投式起子培养物是直接添加到桶中的高度浓缩的生物质(通常为1010到1012个cfu/g)。优点包括减少了噬菌体攻击的风险、使用的灵活性、可获得混合菌株和物种培养物以及不需要繁殖设施。冷冻干燥培养物通常储存-18℃，但冷冻培养物在运输期间需要用干冰冷却并储存在-45℃。
用于起子培养物的典型生产工艺包括以下步骤：(a)处置接种材料，(b)制备培养基，(c)在pH控制下在发酵器中繁殖培养物，(d)浓缩，(e)冷冻，(f)干燥以及(g)包装和储存。生产起子培养物的步骤对于获得培养物产品的期望特性、纯度和质量是重要的。在无菌条件下制备用作直接接种材料的培养物，并将转移保持在最低限度。
用于生产培养物的生长培养基可含有所选乳组分并补充各种营养素，诸如酵母提取物、维生素和矿物质。将培养物生长培养基加热到超高温，并且对于嗜中温培养物或嗜热培养物，分别冷却到30或40℃。在接种培养物后，通过添加碱(诸如NaOH或NH4OH)将pH值维持在6.0-6.3(对于嗜中温培养物)和5.5-6.0(对于嗜热培养物)来优化生长。
对处理参数(诸如发酵器中的温度、搅动速率和顶空气体)进行调整，以产生比批量起子浓缩得多的细胞悬浮液。在发酵(通常是容量为10,000到40,000L的容器中的分批发酵)后，将内容物冷却并收获生物质，使细胞进一步浓缩10-20倍。通常采用分离装置来分离水性液体以收集细菌。
由于成本效益和效率，因此不断需要更高的生物质产率。对于DVS用途，期望使细菌培养物尽可能浓缩，同时具有尽可能高的细胞回收率。
为了包含足够的细菌，商业相关的高度浓缩培养物通常具有高水平的锰。本领域已知锰促进细菌、特别是乳酸菌的生长。对于生物质产率是首要关注的起子培养物制造商来说，这是重要的经济考虑因素(Raccach，M.“锰和乳酸菌(Manganese and lactic acidbacteria).”Journal of food protection 48.10(1985):895-898)。
EP2119766公开了锰可增加乳酸菌的生长产率。
EP0130228公开了锰盐可用于乳酸菌的快速发酵。已常规地以例如食品级锰盐的形式，以适当的量添加锰以促进细胞生长。
最终的浓缩培养物经常含有高水平的锰。可使用的食品级锰盐的示例包括氯化锰、氧化锰、硫酸锰、柠檬酸锰、甘油磷酸锰、葡萄糖酸锰等。锰盐可在接种细菌之前或者与所述接种同时添加到发酵培养基中。
本发明大体上涉及包含乳酸菌的组合物，所述组合物用于添加到诸如食物产品的产品中。所述细菌组合物可用作用于食物产品的起子培养物组合物。其可被添加到食物产品中，或者被添加以发酵食物产品并同时管理真菌生长。所述组合物的特征在于其含有低水平或降低水平的锰。如本上下文中使用的术语“起子培养物”是指一种或多种能够酸化食物产品的细菌的培养物。
在第一方面，本申请提供包含乳酸菌和低水平的锰(诸如至多600ppm的锰)的起子培养物组合物。所述组合物可用于摄取产品中反之可供酵母或霉菌利用的游离锰。本发明人已经证明，常见的酵母和霉菌被所公开的组合物抑制。
Archibald等(1984)探索了植物乳杆菌(L.plantarum)14917对锰的摄取(Archibald等，“植物乳杆菌对锰的获取(Manganese acquisition by Lactobacillusplantarum).”Journal of bacteriology 158.1(1984):1-8)。如所公开的，植物乳杆菌14917的高锰含量是由特定的高亲和力、高速摄取系统生成的。然而，这项工作不涉及食物产品中污染酵母和/或霉菌的生长抑制。它也没有公开高度浓缩的直投式起子培养物(诸如冷冻直投式(F-DVS)或冷冻干燥直投式(FD-DVS))的任何制备。
在一个方面，本发明提供包含一种或多种抗真菌乳酸菌的直投式起子培养物组合物，其用于发酵食物产品和用于抑制或延迟所述食物产品中的真菌生长，其特征在于，所述组合物包含至多600ppm的锰并且乳酸菌的浓度为至少1E+10个菌落形成单位/g。
在一些实施方案中，本文提供包含具有锰转运蛋白的乳酸菌的冷冻直投式(F-DVS)或冷冻干燥直投式(FD-DVS)起子培养物组合物，其用于发酵食物产品和用于抑制或延迟所述食物产品中的真菌生长，其特征在于，所述组合物包含至多600ppm的锰并且其中乳酸菌的浓度为至少1E+10个菌落形成单位/g。
优选地，起子培养物组合物中的细菌浓度为至少1E+10个菌落形成单位(CFU)/g。
为了应对微生物腐败问题，本发明在其他方面提供用于抑制或延迟食物产品中真菌生长的具有至少1E+10个CFU/g的细菌起子组合物，其特征在于，所述组合物包含至多600ppm的锰。优选地，细菌组合物是冷冻干燥直投式(FD-DVS)或冷冻直投式(F-DVS)。所述组合物优选包含一种或多种乳酸菌菌株。
本文还提供用于抑制或延迟基于乳的产品中的真菌生长的起子培养物组合物或用于发酵基于乳的食物产品和抑制或延迟所述食物产品中的真菌生长的起子培养物组合物，所述组合物包含乳酸菌，其特征在于，所述组合物包含至多600ppm的锰并且其中乳酸菌菌落形成单位/g的浓度为至少1E+10，优选其中乳酸菌使乳、酒、茶、植物和/或肉基质发酵。
所述组合物、优选冷冻干燥的DVS(FD-DVS)或冷冻的DVS(F-DVS)可包含至多550ppm的锰、至多500ppm的锰、至多450ppm的锰、至多400ppm的锰、至多350ppm的锰、至多300ppm的锰、至多250ppm的锰、至多200ppm的锰、至多150ppm的锰、至多100ppm的锰、至多70ppm的锰、至多50ppm的锰、至多40ppm的锰。
所述组合物可包含10-600ppm的锰、30-600ppm的锰、35-600ppm的锰、40-600ppm的锰、45-600ppm的锰、50-600ppm的锰、60-550ppm的锰、100-500ppm的锰、150-450ppm的锰、190-400ppm的锰、200-350ppm的锰、250-300ppm的锰。优选地，所述组合物可包含40-250ppm的锰，更优选地，所述组合物可包含45-200ppm的锰。
所述组合物可包含具有1E+10-5E+12、优选2.0E+10-6.5E+11、更优选6.0E+10-6.4E+11、甚至更优选1.3E+11-5.6E+11的菌落形成单位/g的细胞的乳酸菌。
在优选的实施方案中，现在所公开的组合物可以是冷冻干燥直投式(FD-DVS)或冷冻直投式(F-DVS)，特别地，所述组合物可以是含有无锰或基本上无锰的添加剂或冷冻保护剂的冷冻干燥直投式(FD-DVS)，或者所述组合物可以是含有无锰或基本上无锰的添加剂或冷冻保护剂的冷冻直投式(F-DVS)。优选地，所述组合物是冷冻干燥直投式(FD-DVS)，并且其可进一步包含选自以下的添加剂(或冷冻保护剂)：酪蛋白酸钠、肌醇、谷氨酸单钠、抗坏血酸钠、蔗糖、麦芽糖糊精、肌苷一磷酸(IMP)、肌苷、聚山梨糖醇酯80、谷氨酸、赖氨酸、谷氨酸钠、麦芽提取物、乳清粉、酵母提取物、麸质、胶原蛋白、明胶、弹性蛋白、角蛋白、白蛋白、碳水化合物或其混合物。优选地，添加剂(或冷冻保护剂)不含锰或基本上不含锰。
所述组合物另外可含有冷冻保护剂和/或常规添加剂作为其他组分，包括营养素，诸如酵母提取物、糖和维生素，例如维生素A、C、D、K或B族维生素。可添加到本发明组合物中的合适冷冻保护剂是改善微生物的耐寒性的组分，诸如甘露醇、山梨糖醇、三聚磷酸钠、木糖醇、甘油、棉子糖、麦芽糖糊精、赤藓醇、苏糖醇、海藻糖、葡萄糖和果糖。其他添加剂可包括例如碳水化合物、调味剂、矿物质、酶(例如粗制凝乳酶(rennet)、乳糖酶和/或磷脂酶)。
在实施方案中，所述组合物可以是含有无锰或基本上无锰的添加剂或冷冻保护剂的冷冻直投式(F-DVS)，所述冷冻保护剂的浓度为每重量F-DVS形式10-40重量％的冷冻保护剂，或每重量F-DVS形式20-35重量％的冷冻保护剂，诸如每重量F-DVS形式31重量％的冷冻保护剂。
在优选的实施方案中，所述组合物是包含碳水化合物作为添加剂的冷冻干燥直投式(FD-DVS)，优选其中添加剂(或冷冻保护剂)不含锰。合适的示例包括选自由以下组成的组的碳水化合物：戊糖(例如核糖、木糖)、己糖(例如果糖、甘露糖、山梨糖)、二糖(例如ducrose、drehalose、蜜二糖、乳果糖)、低聚糖(例如棉子糖)、低聚果糖(例如actilight、fribrolose)、多糖(例如麦芽糖糊精、黄原胶、果胶、海藻酸盐、微晶纤维素、右旋糖酐、聚乙二醇和糖醇(山梨糖醇、甘露醇)。优选的碳水化合物是二糖，优选海藻糖、蔗糖和/或麦芽糖糊精。
在优选的实施方案中，当处于冷冻状态时，所述组合物可包含以每重量DVS形式的添加剂重量测量的2％到70％的添加剂(或冷冻保护剂)，更优选以每重量DVS形式的添加剂重量测量的3％到50％的添加剂(或冷冻保护剂)，甚至更优选以每重量DVS形式的添加剂重量测量的4％到40％的添加剂(或冷冻保护剂)，且最优选以每重量DVS形式的添加剂重量测量的10％到35％的添加剂(或冷冻保护剂)。优选地，添加剂不含锰或基本上不含锰。
在本发明的上下文中，当所述添加剂或冷冻保护剂中存在小于10ppm的浓度的锰时，所述添加剂或冷冻保护剂不含锰或基本上不含锰。此外，在本发明的上下文中，“添加剂”、“冷冻保护剂(cryoprotectant)”或“冷冻保护试剂(cryoprotectant agent)”可互换使用。
可通过在合适的温度将固体冷冻保护剂与生物质混合例如30分钟来将添加剂(或冷冻保护剂)添加到发酵后的分离的活细菌(生物质)中。如果冷冻保护剂是例如蔗糖，则合适的温度可以是室温。或者，可将添加剂(或冷冻保护剂)的无菌溶液与生物质混合。对于蔗糖，合适的无菌溶液可由50％(w/w)蔗糖溶液制成。对于海藻糖，合适的无菌溶液可由40％(w/w)溶液制成。
在优选的实施方案中，所述组合物是冷冻干燥的DVS，其包含10-600ppm的锰、30-600ppm的锰、35-600ppm的锰、40-600ppm的锰、45-600ppm的锰、50-600ppm的锰、60-550ppm的锰、100-500ppm的锰、150-450ppm的锰、190-400ppm的锰、200-350ppm的锰、250-300ppm的锰。优选地，所述组合物是包含40-250ppm的锰的冷冻干燥的DVS，更优选地，所述组合物是包含45-200ppm的锰的冷冻干燥的DVS。此外，所述优选的冷冻干燥DVS组合物可包含选自由以下组成的组的乳酸菌：植物乳杆菌、发酵乳杆菌(Lactobacillus fermentum)、罗伊氏乳杆菌(Lactobacillus reuteri)、沙克乳杆菌(Lactobacillus sakei)、短乳杆菌(Lactobacillus brevis)、干酪乳杆菌(Lactobacillus casei)、副干酪乳杆菌(Lactobacillus paracasei)、唾液乳杆菌(Lactobacillus salivarius)、消化乳杆菌(Lactobacillus alimentarius)、乳酸片球菌(Pediococcus acidilactici)、鼠李糖乳杆菌(Lactobacillus rhamnosus)和开菲尔乳杆菌(Lactobacillus kefiri)，优选选自副干酪乳杆菌和/或鼠李糖乳杆菌，其中所述乳酸菌具有至少1E+10的CFU/g，包括2E+10、3E+10、4E+10、5E+10、6E+10、7E+10、8E+10、9E+10 CFU/g，诸如2.0E+10-6.5E+11，优选6.0E+10-6.4E+11，更优选1.3E+11-5.6E+11，和/或其中冷冻干燥DVS具有不含锰或基本上不含锰且浓度为每重量FD-DVS形式10-30％干重的冷冻保护剂或每重量FD-DVS形式20-30％干重的冷冻保护剂(诸如每重量FD-DVS形式27％干重的冷冻保护剂)的添加剂或冷冻保护剂。
在优选的实施方案中，本发明可提供用于抑制或延迟食物产品中真菌生长的组合物、或用于抑制或延迟基于乳的产品中真菌生长的组合物、或用于发酵基于乳的食物产品以及抑制或延迟所述食物产品中真菌生长的组合物，其中所述乳酸菌包含锰转运蛋白，所述锰转运蛋白与SEQ ID NO:1-3中任一个的序列具有至少55％，诸如至少60％、65％、70％、75％、80％、85％、90％、95％、96％、97％、98％、99％或100％的序列同一性。
在优选的实施方案中，本发明可提供用于抑制或延迟食物产品中真菌生长的组合物、或用于抑制或延迟基于乳的产品中真菌生长的组合物、或用于发酵基于乳的食物产品以及抑制或延迟所述食物产品中真菌生长的组合物，其中乳酸菌不含超氧化物歧化酶，优选不含锰超氧化物歧化酶。
超氧化物歧化酶(诸如锰超氧化物歧化酶)已得到研究，并且例如尤其描述于以下文献中：Kehres等，“细菌中锰转运、生物化学和发病机制的新兴主题(Emerging themes inmanganese transport,biochemistry and pathogenesis in bacteria).”FEMSmicrobiology reviews 27.2-3(2003):263-290；Culotta V.C“超氧化物歧化酶、氧化应激和细胞代谢(Superoxide dismutase,oxidative stress,and cell metabolism)”Curr.Top.Cell Regul.36,117-132(2000)或Whittaker J.W“锰超氧化物歧化酶(Manganese superoxide dismutase)”Met.Ions Biol.Syst.37,587-611(2000)。
在本发明的上下文中，术语“不含”意指一种或多种细菌菌株的基因组不呈现编码超氧化物歧化酶的基因，或者即使一种或多种细菌菌株的基因组呈现编码超氧化物歧化酶的基因，该基因也不被一种或多种细菌菌株表达。
在优选的实施方案中，本发明可提供用于抑制或延迟食物产品中真菌生长的组合物、或用于抑制或延迟基于乳的产品中真菌生长的组合物、或用于发酵基于乳的食物产品以及抑制或延迟所述食物产品中真菌生长的组合物，其中乳酸菌选自由以下组成的组：植物乳杆菌、发酵乳杆菌、罗伊氏乳杆菌、沙克乳杆菌、短乳杆菌、干酪乳杆菌、副干酪乳杆菌、唾液乳杆菌、消化乳杆菌、乳酸片球菌、鼠李糖乳杆菌和开菲尔乳杆菌。
在优选的实施方案中，本发明可提供用于抑制或延迟食物产品中真菌生长的组合物、或用于抑制或延迟基于乳的产品中真菌生长的组合物、或用于发酵基于乳的食物产品以及抑制或延迟所述食物产品中真菌生长的组合物，其中真菌是酵母和/或霉菌，优选其中真菌是选自由以下组成的组的酵母：有孢圆酵母属(Torulaspora)的种、隐球菌属(Cryptococcus)的种、酵母属(Saccharomyces)的种、耶氏酵母属(Yarrowia)的种、德巴利酵母属(Debaryomyces)的种、假丝酵母属(Candida)的种和红酵母属(Rhodoturola)，优选其中德巴利酵母属的种是汉逊德巴利酵母(Debaromyces hansenii)和/或其中真菌是选自由以下组成的组的霉菌：曲霉属(Aspergillus)的种、枝孢菌属(Cladosporium)的种、亚隔孢壳属(Didymella)的种或青霉属(Penicillium)的种，优选其中青霉属的种是皮壳青霉(Penicillium crustosum)、潘氏青霉(Penicillium paneum)、卡尼青霉(Penicilliumcarneum)或娄地青霉(Penicillium roqueforti)。
在又一方面，本发明还提供包含本文所公开的组合物的食物产品。在实施方案中，食物产品可以是发酵食物产品，优选嗜热发酵食物产品或嗜中温发酵食物产品，更优选所述食物产品可以是酸奶或奶酪。
在第三方面，本发明还提供本文所公开的组合物作为食物产品中真菌生长抑制剂的用途，优选其中食物产品是发酵食物产品，更优选嗜热发酵食物产品或嗜中温发酵食物产品，更优选酸奶或奶酪。
附图说明
图1：用起子培养物(参考)生产的酸奶(1.5％脂肪)、或用起子培养物和鼠李糖乳杆菌菌株1(1E+7个CFU/g)的F-DVS形式(其中所述F-DVS形式具有约30ppm的锰、或约195ppm的锰、或约625ppm的锰)生产的酸奶(1.5％脂肪)、或用起子培养物以及副干酪乳杆菌和鼠李糖乳杆菌菌株3的FD-DVS形式(其中FD-DVS形式的所述形式具有约275ppm的锰)生产的酸奶(1.5％脂肪)。酸奶被掺入皮壳青霉(X)、卡尼青霉(Y)和娄地青霉(Z)(500个孢子/每种)并在22℃储存7天。
图2：当使用50个CFU/g的德氏有孢圆酵母接种用起子培养物(参考)生产的酸奶(1.5％脂肪)、或用起子培养物和鼠李糖乳杆菌菌株1(1E+7个CFU/g)的F-DVS形式(其中所述F-DVS形式具有约30ppm的锰、或约195ppm的锰、或约625ppm的锰)生产的酸奶(1.5％脂肪)时，德氏有孢圆酵母的生长。将酸奶在7℃储存23天。
图3：当使用50个CFU/g的汉逊德巴利酵母接种用起子培养物(参考)生产的酸奶(1.5％脂肪)、或用起子培养物和鼠李糖乳杆菌菌株1(1E+7个CFU/g)的F-DVS形式(其中所述F-DVS形式具有约30ppm的锰、或约195ppm的锰、或约625ppm的锰)生产的酸奶(1.5％脂肪)时，汉逊德巴利酵母的生长。将酸奶在7℃储存23天。
图4：当使用50个CFU/g的汉逊德巴利酵母接种用起子培养物(参考)生产的酸奶(1.5％脂肪)、或用起子培养物和鼠李糖乳杆菌菌株2的F-DVS形式(其中所述F-DVS形式具有约45ppm的锰)生产的酸奶(1.5％脂肪)、或用起子培养物以及鼠李糖乳杆菌菌株1和2的F-DVS形式(其中F-DVS形式具有约65ppm的锰)生产的酸奶(1.5％脂肪)、或用起子培养物和具有约845ppm的锰的基准组合物A的FD-DVS形式生产的酸奶(1.5％脂肪)、或用起子培养物和具有约630ppm的锰的基准组合物B的FD-DVS形式生产的酸奶(1.5％脂肪)、或用起子培养物和具有870ppm的锰的基准组合物C的FD-DVS形式生产的酸奶(1.5％脂肪)时，汉逊德巴利酵母的生长。将酸奶在7℃储存32天。
图5：用起子培养物(参考)生产的酸奶(1.5％脂肪)、或用起子培养物和鼠李糖乳杆菌菌株2(1E+7个CFU/g)的F-DVS形式(其中F-DVS形式具有约45ppm的锰)、或和鼠李糖乳杆菌菌株1和2(1E+7个CFU/g)的F-DVS形式(其中F-DVS形式具有约65ppm的锰)、或和具有约845ppm的锰的基准组合物A(1E+7个CFU/g)的FD-DVS形式、或和具有约630ppm的锰的基准组合物B(1E+7个CFU/g)的FD-DVS形式、或和具有约870ppm的锰的基准组合物C(1E+7个CFU/g)的FD-DVS形式生产的酸奶(1.5％脂肪)。所述酸奶已掺入卡尼青霉、潘氏青霉和娄地青霉(500个孢子/每种)并在7℃储存24天(上行)或在25℃储存6天(下行)。平板上青霉属种的布置与图1相同，只是卡尼青霉被潘氏青霉替代。
图6：当使用50个CFU/g的汉逊德巴利酵母接种用起子培养物(参考)生产的酸奶(1.5％脂肪)、或用起子培养物以及副干酪乳杆菌和鼠李糖乳杆菌菌株3的FD-DVS形式(其中FD-DVS形式的所述形式具有约275ppm的锰)生产的酸奶(1.5％脂肪)、或用起子培养物和鼠李糖乳杆菌菌株2(1E+7个CFU/g)的FD-DVS形式生产的酸奶(1.5％脂肪)(其中所述FD-DVS形式具有约200ppm的锰并且向其中将脱脂乳粉(SMP)添加或不添加到用于获得FD-DVS形式的冷冻保护剂(标准cryo)中)时，汉逊德巴利酵母的生长。将酸奶在7℃储存27天。
图7：用起子培养物和鼠李糖乳杆菌菌株2(1E+7 CFU/g)的FD-DVS形式(其中所述FD-DVS形式具有约200ppm的锰并且向其中将不同浓度的锰(1、5、10、20和40ppm)添加到用于获得FD-DVS形式的冷冻保护剂中)生产的酸奶(1.5％脂肪)。酸奶被掺入皮壳青霉、卡尼青霉和娄地青霉(500个孢子/每种)并在22℃储存12天。平板上青霉属种的布置与图1相同。
图8：当使用50个CFU/g的汉逊德巴利酵母接种用起子培养物和鼠李糖乳杆菌菌株2(1E+7 CFU/g)的FD-DVS形式(其中所述FD-DVS形式具有约200ppm的锰并且向其中将不同浓度的锰(1和40ppm)添加到用于获得FD-DVS形式的冷冻保护剂中)生产的酸奶(1.5％脂肪)时，汉逊德巴利酵母的生长。将酸奶在7℃储存27天。
具体实施方式
食物损失是全球关注的主要问题——为人消耗而生产的所有食物中大约有三分之一损失或浪费。这种全球食物大量损失的原因多种多样，但影响感官产品质量(外观、质地、味道和香气)的微生物腐败起着主要作用。由于真菌可在不同、甚至恶劣的环境中生长，因此它们是在食品加工链所有阶段发现的主要腐败微生物。因此，通过控制真菌污染来减少粮食损失至关重要。
响应于这一要求，本发明提供用于抑制或延迟食物产品中真菌生长的新型组合物。锰在自然界和我们的许多消费品中以痕量存在。最近发现低水平的游离锰浓度可用作用于酵母和/或霉菌生长的限制因素。因此，通过操控给定产品中游离锰的浓度，可有效地管理微生物腐败。此类腐败预防策略甚至适用于食物产品以外的产品，并延伸到通常易于受到微生物污染的其他产品，诸如饲料产品、生物产品、健康护理产品、药物产品等。
许多细菌已经发展出复杂的采集系统来从环境中清除必需金属。因此可应用锰清除细菌来摄取产品中的游离锰。此外，本发明人最近已发现，当在按比例放大工艺中制备细菌时，优选较低水平的锰。
在第一方面，本发明提供用于抑制或延迟基于乳的产品中的真菌生长的起子培养物组合物或用于发酵基于乳的食物产品和抑制或延迟所述食物产品中的真菌生长的起子培养物组合物，所述组合物包含乳酸菌，其特征在于，所述组合物包含至多600ppm的锰并且其中乳酸菌菌落形成单位/g的浓度为至少1E+10，优选其中乳酸菌使乳、酒、茶、植物和/或肉基质发酵。
一般来说，抑制意指细胞或微生物的功能和活性的降低(无论是部分还是全部)。如本文所用，与酵母和霉菌有关的术语“抑制(to inhibit)”和“抑制(inhibiting)”意指酵母和霉菌的生长、数量或浓度相同或降低。这可以通过微生物学领域中已知的任何方法来测量。可通过将产品中或产品上的真菌生长、数量或浓度与对照进行比较来观察抑制。对照可以是相同的产品，但不含所述组合物。
一般来说，术语“延迟”是指停止、推迟、阻碍或引起某事比正常情况更慢地发生的作用。如本文所用，“延迟真菌生长”是指推迟真菌生长的作用。这可通过比较两种产品中真菌生长到给定水平所需的时间来观察，所述产品中的一种具有所公开的组合物而另一种没有。
在一些实施方案中，“抑制或延迟真菌生长”是指延迟7天，诸如8、9、10、11、12、13、14、15、16、17、18、20、21、22、23、24、25、30、35、40、45、50、55、60天。
术语“抗真菌”在本申请中应理解为抑制延迟食物产品(诸如基于乳的产品)中真菌生长的能力。
如本文所用，术语“食物基质”是指食物的组成和结构。它基于营养素包含在连续介质中的概念。
如本文所用，术语“肉基质”是指肉的组成和结构。它基于营养素包含在连续介质中的概念。
锰
锰参与许多至关重要的生物过程，并且普遍见于所有生物体中。锰还有助于防止氧化应激，并且还有助于活性氧物质的催化解毒。许多细菌已经发展出复杂的采集系统来使用用于螯合或游离金属的低亲和力和高亲和力转运系统从环境中清除必需金属。被细菌摄取的锰在蛋白质中形成大的不可透析的聚磷酸盐-蛋白质聚集体复合物，其可能达到非常高的细胞内浓度。
根据本申请的“锰”是指存在于组合物中用于抑制或延迟食物产品中真菌生长的锰。在本发明的上下文中，“锰”包括见于细胞内和细胞外的锰。
如本文所用，术语“细菌菌株”具有在微生物学领域的普通含义并且是指细菌的遗传变体。
如本文所用，锰浓度或锰水平以基于重量/重量计算的百万分率(“ppm”)表示。产品或组合物中锰的浓度低于某一值意味着产品或其部分中锰的浓度使得按重量计整个产品或整个组合物中锰的浓度低于给定值。确定痕量元素(诸如锰)的方法是本领域已知的并且描述于例如Nielsen,S.Suzanne编辑，Food analysis.第86卷，Gaithersburg,MD:AspenPublishers,1998中。
测量低浓度锰的方法为本领域技术人员所熟知。此类方法包括原子吸收光谱、原子发射光谱、质谱、中子活化分析和x射线荧光测定(参见例如Williams等“锰的毒理学特征(Toxicological profile for manganese).”(2012))。
优选地，根据由欧洲标准化委员会(European Committee for Standardization)出版的欧洲标准EN13805:2014中的“食品-痕量元素的确定-加压溶出(Foodstuffs-Determination of trace elements-Pressure digestion)”中所述的标准程序、或如由国际标准化组织(International Organization for Standardization)出版的ISO 11885:2007中的“水质-通过电感耦合等离子体光发射光谱确定所选元素(Water quality-Determination of selected elements by inductively coupled plasma opticalemission spectrometry)(ICP-OES)”中所述的标准程序测量锰浓度。
存在于最终组合物(F-DVS产品或FD-DVS产品)中的锰水平被确定为影响菌株抗真菌活性的主要参数之一，其中高水平的Mn产生较小的抗真菌活性且低水平的Mn产生高的抗真菌活性。
先前已经研究了来自植物乳杆菌的镉和锰摄取基因。在该研究中，Hao等公开了植物乳杆菌ATCC14917中的两种镉摄取系统。一种与Mn2+不足无关，但具有低亲和力，而另一种具有高亲和力且由Mn2+不足诱导，但在Mn2+存在下受到抑制。对于后者，Mn2+和Cd2+是彼此的竞争性抑制剂，且其对Cd2+的亲和力高于对Mn2+的亲和力(Hao等“来自植物乳杆菌的镉和锰摄取基因的克隆、表达及表征(Cloning,expression,and characterization of cadmiumand manganese uptake genes from Lactobacillus plantarum).”Applied andEnvironmental Microbiology 65.11(1999):4746-4752和Hao等“植物乳杆菌中镉摄取的表征以及镉和锰摄取突变体的分离(Characterization of cadmium uptake inLactobacillus plantarum and isolation of cadmium and manganese uptakemutants).”Applied and environmental microbiology 65.11(1999):4741-4745.)。这些论文不涉及用于直接接种的高生物量细胞培养物。它们既没有教导也没有提示本申请的发现。
本领域技术人员能够调整培养基中的锰水平以获得含有期望锰水平的最终产品。例如，如果生长培养基中的锰水平低，则最终的组合物将因此具有低水平的锰，因为锰预计会保留在浓缩过程中。另一方面，如果生长培养基中的锰水平高，则最终的组合物将相应地具有高水平的锰。
真菌
真菌是属于真菌界的成员。可用本领域技术人员已知的各种方法测量真菌生长。例如，真菌生长可通过菌落的密度或大小、细胞数量、菌丝团变化、孢子产生、菌丝生长、菌落形成单位(CFU)等来测量，这取决于真菌类型和应用所述方法的产品。还可通过测量营养素或代谢物浓度的变化(诸如二氧化碳释放和氧气摄取)来观察真菌生长。
术语“真菌生长的抑制”或“抑制真菌生长”是指真菌细胞增殖的抑制。
术语“真菌生长的延迟”或“延迟真菌生长”是指真菌细胞增殖的减缓。这可例如通过测量真菌生长并将其与对照进行比较来观察。此类对照可以是例如在没有现在所公开的组合物的情况下制备的产品。确定真菌生长抑制或延迟的方法是本领域技术人员已知的。
在一个实施方案中，现在所公开的组合物抑制诸如以下的酵母的生长：假丝酵母属的种、迈耶氏酵母属(Meyerozyma)的种、克鲁维酵母属(Kluyveromyces)的种、毕赤酵母属(Pichia)的种、半乳糖酵母属(Galactomyces)的种、毛孢子菌属(Trichosporon)的种、锁掷酵母属(Sporidiobolus)的种、有孢圆酵母属的种、隐球菌属(Cryptococcus)的种、酵母属的种、耶氏酵母属的种、德巴利酵母属的种和红酵母属的种。优选地，真菌是选自由以下组成的组的酵母：有孢圆酵母属的种、隐球菌属的种、酵母属的种、耶氏酵母属的种、德巴利酵母属的种、假丝酵母属的种和红酵母属的种。更优选地，真菌是选自由以下组成的组的酵母：德氏有孢圆酵母、长莓隐球酵母(Cryptococcus fragicola)、酿酒酵母(Sacharomycescerevisiae)、解脂耶氏酵母(Yarrowia lipolytica)、汉逊德巴利酵母和胶红酵母(Rhodoturola mucilaginosa)。
在一个实施方案中，现在所公开的组合物抑制霉菌的生长。优选地，真菌是选自由以下组成的组的霉菌：曲霉属的种、枝孢菌属的种、亚隔孢壳属的种或青霉属的种。更优选地，真菌是选自由以下组成的组的霉菌：短密青霉(Penicillium brevicompactum)、皮壳青霉、离生青霉(Penicillium solitum)、卡尼青霉、潘氏青霉和娄地青霉。
乳酸菌(LAB)
“乳酸菌”指使糖发酵同时产生酸(包括作为主要产生酸的乳酸)的革兰氏阳性微需氧或厌氧细菌。食物产品通常具有约3.5到约6.5(诸如约4到约6，诸如约4.5到约5.5，诸如约5)的pH值。
在优选的实施方案中，现在所公开的组合物可包含具有用于锰的转运系统的乳酸菌。这些用于锰的转运系统已得到研究，并且例如描述于Kehres等，“细菌中锰转运、生物化学和发病机制的新兴主题.”FEMS microbiology reviews 27.2-3(2003):263-290中。
可用于本申请的乳酸菌菌株具有锰摄取活性。通过常规实验，本领域技术人员能够选择具有锰摄取活性的细菌。此类细菌可例如包含细菌Mn2+转运蛋白。Mn2+转运蛋白可以是ABC转运蛋白(例如SitABCD和YfeABCD)或质子依赖性Nramp相关转运系统，该转运系统属于在由转运分类数据库(Transport Classification Database)给出的转运蛋白分类系统中指定为TC#3.A.1.15和TC#2.A.55的家族(M.Saier；U of CA,San Diego,Saier MH,ReddyVS,Tamang DG,Vastermark A.(2014))。TC系统是用于转运蛋白的分类系统，其类似于用于酶分类的酶委员会(Enzyme Commission,EC)系统。转运蛋白分类(TC)系统是由国际生物化学和分子生物学联盟(International Union of Biochemistry and Molecular Biology)批准的转运蛋白分类命名系统。TCDB可在http://www.tcdb.org免费访问，该网站提供了几种不同的用于访问数据的方法，包括逐步访问分级分类、按序列或TC编号直接检索和全文检索。
在优选的实施方案中，乳酸菌菌株可包含属于指定为TC#3.A.1.15(锰螯合物摄取转运蛋白(MZT)家族)或TC#3.A.1.15.6、TC#3.A.1.15.8、TC#3.A.1.15.14的家族的蛋白质或其功能变体。
ABC转运蛋白主要在较高的pH值下具有活性，而质子驱动的转运蛋白在酸性条件下可能更具活性。因此，在一个实施方案中，现在所公开的组合物可包含细菌菌株，所述细菌菌株包含属于指定为TC#2.A.55家族(金属离子(Mn2+-铁)转运蛋白(Nramp)家族)的蛋白质或其功能变体。更优选地，转运蛋白属于指定为TC#2.A.55.2的亚族或指定为TC#2.A.55.3的亚族。
例如，本文所公开的组合物可包含乳酸菌，所述乳酸菌具有指定为TC#2.A.55.3.1、TC#2.A.55.3.2、TC#2.A.55.3.2、TC#2.A.55.3.3、TC#2.A.55.3.4、TC#2.A.55.3.5、TC#2.A.55.3.6、TC#2.A.55.3.7、TC#2.A.55.3.8或TC#2.A.55.3.9金属离子(Mn2+-铁)转运蛋白或其功能变体，优选具有TC#2.A.55.2.6或其功能变体。
术语“功能变体”是具有基本上相似的生物活性(即锰摄取活性)的蛋白质变体。
如本文所用，“变体”是指蛋白质的变体形式，其与该蛋白质的特定核酸或氨基酸序列共有至少55％、60％、65％、70％、75％、80％、85％、90％、95％、96％、97％、98％、99％或100％序列同一性。
本公开另外提供可存在于乳酸中的锰转运蛋白的多肽序列以实施本发明。
在优选的实施方案中，乳酸菌菌株包含具有SEQ ID NO:1(MASEDKKSKREHIIHFEDTPSKSLDEVNGSVEVPHNAGFWKTLAAYTGPGILVAVGYMDPGNWITSIAGGASFKYSLLSVILISSLIAMLLQAMAARLGIVTGRDLAQMTRDHTSKAMGGFLWVITELAIMATDIAEIIGSAIALKLLFNMPLIVGIIITTADVLILLLLMRLGFRKIEAVVATLVLVILLVFAYEVILAQPNVPELLKGYLPHADIVTNKSMLYLSLGIVGATVMPHDLFLGSSISQTRKIDRTKHEEVKKAIKFSTIDSNLQLTMAFIVNSLLLILGAALFFGTSSSVGRFVDLFNALSNSQIVGAIASPMLSMLFAVALLASGQSSTITGTLAGQIIMEGFIHLKMPLWAQRLLTRLMSVTPVLIFAIYYHGNEAKIENLLTFSQVFLSIALPFAVIPLVLYTSDKKIMGEFANRAWVKWTAWFISGVLIILNLYLIAQTLGFVK)的序列的多肽或其功能变体。
在其他优选的实施方案中，乳酸菌菌株包含与SEQ ID NO:1的序列具有至少55％(诸如至少60％、65％、70％、75％、80％、85％、90％、95％、96％、97％、98％、99％或100％)序列同一性的多肽。
表1显示编码SEQ ID NO:1的功能变体的示例性序列和它们与SEQ ID NO:1的序列同一性.
表1
Figure BDA0003604753450000191
Figure BDA0003604753450000201
Figure BDA0003604753450000211
Figure BDA0003604753450000221
在优选的实施方案中，乳酸菌菌株包含具有SEQ ID NO:2(MARPDERLTVQREKRSLDDINRSVQVPSVYESSFFQKFLAYSGPGALVAVGYMDPGNWLTALEGGSRYHYALLSVLLMSILVAMFMQTLAIKLGVVARLDLAQAIAAFIPNWSRICLWLINEAAMMATDMTGVVGTAIALKLLFGLPLMWGMLLTIADVLVVLLFLRFGIRRIELIVLVSILTVGIIFGIEVARADPSIGGIAGGFVPHTDILTNHGMLLLSLGIMGATIMPHNIYLHSSLAQSRKYDEHIPAQVTEALRFGKWDSNVHLVAAFLINALLLILGAALFYGVGGHVTAFQGAYNGLKNPMIVGGLASPLMSTLFAFALLITGLISSIASTLAGQIVMEGYLNIRMPLWERRLLTRLVTLIPIMVIGFMIGFSEHNFEQVIVYAQVSLSIALPFTLFPLVALTNRRDLMGIHVNSQLVRWVGYFLTGVITVLNIQLAISVFV)的序列的多肽或其功能变体。
在其他优选的实施方案中，乳酸菌菌株包含与SEQ ID NO:2的序列具有至少55％(诸如至少60％、65％、70％、75％、80％、85％、90％、95％、96％、97％、98％、99％或100％)序列同一性的多肽。
表2显示编码SEQ ID NO:2的功能变体的示例性序列和它们与SEQ ID NO:2的序列同一性.
表2
Figure BDA0003604753450000222
Figure BDA0003604753450000231
Figure BDA0003604753450000241
在优选的实施方案中，乳酸菌菌株包含具有SEQ ID NO:3(MSDDHKKRHPIKLIQYANGPSLEEINGTVEVPHGKGFWRTLFAYSGPGALVAVGYMDPGNWSTSITGGQNFQYLLISVILMSSLIAMLLQYMAAKLGIVSQMDLAQAIRARTSKKLGIVLWILTELAIMATDIAEVIGAAIALYLLFHIPLVIAVLVTVLDVLVLLLLTKIGFRKIEAIVVALILVILLVFVYQVALSDPNMGALLKGFIPTGETFASSPSINGMSPIQGALGIIGATVMPHNLYLHSAISQTRKIDYKNPDDVAQAVKFSAWDSNIQLSFAFVVNCLLLVMGVAVFKSGAVKDPSFFGLFQALSDSSTLSNGVLIAVAKSGILSILFAVALLASGQNSTITGTLTGQVIMEGFVHMKMPLWARRLVTRIISVIPVIVCVMLTARDTPIQQHEALNTLMNNSQVFLAFALPFSMLPLLMFTNSKVEMGDRFKNTGWVKVLGWISVLGLTGLNLKGLPDSIAGFFGDHPTATQTNMANIIAIVLIVAILALLAWTIWDLYKGNQRYEAHLAAVADEKEAKADVDEQ)的序列的多肽或其功能变体。
在其他优选的实施方案中，乳酸菌菌株是包含与SEQ ID NO:3的序列具有至少55％(诸如至少60％、65％、70％、75％、80％、85％、90％、95％、96％、97％、98％、99％或100％)序列同一性的多肽的细菌菌株。
表3显示编码SEQ ID NO:3的功能变体的示例性序列和它们与SEQ ID NO:3的序列同一性.
表3
Figure BDA0003604753450000251
Figure BDA0003604753450000261
Figure BDA0003604753450000271
出于本发明的目的，两个氨基酸序列之间的“序列同一性”程度是使用如在EMBOSS软件包(EMBOSS:The European Molecular Biology Open Software Suite，Rice等，2000，Trends Genet.16:276-277)(优选版本3.0.0或更新的版本)的Needle程序中执行的Needleman-Wunsch算法(Needleman和Wunsch,1970,J.Mol.Biol.48:443-453)来确定的。所用的任选参数是10的缺口开放罚分(gap open penalty)、0.5的缺口延伸罚分(gapextension penalty)和EBLOSUM62(BLOSUM62的EMBOSS版本)取代矩阵(substitutionmatrix)。使用标记为“最长同一性(longest identity)”(使用nobrief选项获得)的Needle输出作为同一性百分比，并计算如下：
(相同的残基×100)/(比对长度-比对中缺口的总数)
出于本发明的目的，两个脱氧核糖核苷酸序列之间的序列同一性程度是使用如在EMBOSS软件包(EMBOSS:The European Molecular Biology Open Software Suite,Rice等，2000，见上文)(优选版本3.0.0或更新的版本)的Needle程序中执行的Needleman-Wunsch算法(Needleman和Wunsch,1970，见上文)来确定的。所用的任选参数是10的缺口开放罚分、0.5的缺口延伸罚分和EDNAFULL(NCBI NUC4.4的EMBOSS版本)取代矩阵。使用标记为“最长同一性”(使用nobrief选项获得)的Needle输出作为同一性百分比，并计算如下：
(相同的脱氧核糖核苷酸×100)/(比对长度-比对中缺口的总数)
在实施方案中，乳酸菌菌株包含与SEQ ID NO:1-3中任一个的序列具有至少55％(诸如至少60％、65％、70％、75％、80％、85％、90％、95％、96％、97％、98％、99％或100％)序列同一性的锰转运蛋白。该确定可基于对细菌菌株进行测序或在已知序列数据库中进行blast搜索。
用于本公开的实施例部分中的乳酸菌具有作为编码的SEQ ID NO:1-3或其功能变体的锰转运蛋白。
在优选的实施方案中，乳酸菌可选自由以下组成的组：植物乳杆菌、发酵乳杆菌、罗伊氏乳杆菌、沙克乳杆菌、短乳杆菌、干酪乳杆菌、副干酪乳杆菌、唾液乳杆菌、消化乳杆菌、乳酸片球菌、鼠李糖乳杆菌和开菲尔乳杆菌。
直投式(DVS)
在一个实施方案中，本发明提供包含乳酸菌的冷冻直投式(F-DVS)起子培养物组合物，其用于发酵食物产品和用于抑制或延迟所述食物产品中的真菌生长，其特征在于，所述组合物包含至多600ppm的锰并且乳酸菌的浓度为至少1E+10个菌落形成单位/g。
在另一实施方案中，本发明提供包含乳酸菌的冷冻干燥直投式(FD-DVS)起子培养物组合物，其用于发酵食物产品和用于抑制或延迟所述食物产品中的真菌生长，其特征在于，所述组合物包含至多600ppm的锰并且乳酸菌的浓度为至少1E+10个菌落形成单位/g。
在一些实施方案中，乳酸菌是植物乳杆菌、发酵乳杆菌、罗伊氏乳杆菌、沙克乳杆菌、短乳杆菌、干酪乳杆菌、副干酪乳杆菌、唾液乳杆菌、消化乳杆菌、乳酸片球菌、鼠李糖乳杆菌和开菲尔乳杆菌。优选地，乳酸菌为副干酪乳杆菌或鼠李糖乳杆菌。
冷冻干燥或冷冻的细菌培养物的制备是本领域已知的，例如如US9848615中所公开的。
商业相关的高浓缩的冷冻培养物通常具有至少1E+9个CFU/g的细菌浓度。在优选的实施方案中，根据本发明的组合物中的细菌具有至少1E+10 CFU/g(包括2E+10、3E+10、4E+10、5E+10、6E+10、7E+10、8E+10、9E+10 CFU/g)的浓度。在其他实施方案中，组合物中的细菌具有至少1E+11 CFU/g(包括2E+10、3E+10、4E+10、5E+10、6E+10、7E+10、8E+10、9E+10个CFU/g)的浓度。为了达到这一目的，可能需要更长的发酵时间。
产品
在一些实施方案中，产品是食物产品、化妆品、健康护理产品或药物产品。“食品”和“食物产品”具有这些术语的共同含义。“食物产品”是指任何适人或动物消耗的食品或饲料产品。食物产品可以是新鲜或易腐烂的食物产品以及储存或加工的食物产品。食物产品包括但不限于水果和蔬菜(包括衍生产品)、谷物和谷物衍生产品、乳制品、肉类、家禽和海产食品。更优选地，食物产品是肉类产品或乳制品，诸如酸奶、特沃劳格(tvarog)、酸奶油、奶酪等。食物产品还可以是基于植物的产品或即食产品，诸如沙拉。
在一个实施方案中，可将本文所述的组合物添加到非发酵食物产品或发酵食物产品中。非发酵产品通常具有高于发酵食物产品的pH值。发酵食物产品是通过微生物的作用产生或保存的食品。发酵意指通过微生物的作用将碳水化合物转化为醇或酸。发酵通常是指使用酵母将糖发酵成酒精。然而，它还可能涉及将乳糖转化为乳酸。例如，发酵可用于制作诸如酸奶、奶酪、萨拉米香肠(salami)、德国泡菜(sauerkraut)、朝鲜泡菜(kimchi)、腌菜(pickle)等食品。
本发明特别可用于抑制或延迟乳制品中的真菌生长。在此类产品中，酵母和霉菌的污染很常见，并限制了此类产品的保存期。除了乳之外，“乳制品”还包括来源于乳的产品，诸如奶油、冰淇淋、黄油、奶酪和酸奶，以及次级产品，诸如抗乳血清和酪蛋白，以及含有乳或乳组分作为主要成分的任何预制食品，诸如配方乳。在一个优选的实施方案中，乳制品是发酵乳制品。术语“乳”被理解为通过给任何哺乳动物(诸如奶牛、绵羊、山羊、水牛或骆驼)挤奶而获得的乳分泌物。在优选的实施方案中，乳是牛乳。术语乳还包括由植物材料制得的蛋白质/脂肪溶液，例如豆乳。
在一个实施方案中，食物产品是通过用嗜热生物发酵制备的产品，即嗜热发酵食物产品。术语“嗜热生物”是指在高于43℃的温度下繁衍最好的微生物。工业上最有用的嗜热菌包括链球菌属的种(Streptococcus spp.)和乳杆菌属的种(Lactobacillus spp.)。术语“嗜热发酵”在本文中是指在高于约35℃(诸如约35℃到约45℃)的温度下的发酵。“嗜热性发酵食物产品”是指通过嗜热性起子培养物的嗜热发酵制备的发酵食物产品。此类产品包括例如酸奶、冰岛酸奶(skyr)、黎巴嫩浓缩酸奶(labneh)、印度酸奶(lassi)、爱兰(ayran)和杜格酸奶(doogh)。
在一个实施方案中，食物产品是通过用嗜中温生物(mesophile)发酵制备的产品，即嗜中温发酵食物产品。术语“嗜中温生物”是指在中等温度(15℃-40℃)下繁衍最好的微生物。工业上最有用的嗜中温菌包括乳球菌属的种(Lactococcus spp.)和明串珠菌属的种(Leuconostoc spp)。术语“嗜中温发酵”在本文中是指在约22℃到约35℃的温度下的发酵。“嗜中温发酵食物产品”是指通过嗜中温起子培养物的嗜中温发酵制备的发酵食物产品。此类产品中包例如酪乳、酸乳、培养乳(cultured milk)、斯美塔那(smetana)、酸奶油和新鲜奶酪，诸如夸克(quark)、特沃劳格和奶油奶酪。
发酵产品的制备
本文的组合物特别可用于抑制或延迟发酵乳产品(诸如嗜热和嗜中温发酵乳产品，例如酸奶产品)中的酵母和/或霉菌生长。术语“发酵乳产品”是一般根据本领域众所周知的相关官方法规和标准定义的术语。例如，将嗜热链球菌(Streptococcusthermophilus)和德氏乳杆菌保加利亚亚种(Lactobacillus delbrueckiisubsp.bulgaricus)的共生培养物用作酸奶的起子培养物，而将嗜酸乳杆菌用于制作嗜酸乳。将其他嗜中温乳酸菌用于生产夸克或清爽奶酪(fromage frais)。
表述“发酵乳产品”是指食品或饲料产品，其中食品或饲料产品的制备涉及用乳酸菌使乳基质发酵。如本文所用的“发酵乳产品”包括但不限于诸如嗜热发酵乳产品(例如酸奶)和嗜中温发酵乳产品(诸如酸奶油和酪乳，以及发酵乳清、夸克和清爽奶酪)的产品。发酵乳产品还包括奶酪，诸如欧式奶酪(continental type cheese)、新鲜奶酪、软奶酪、切达奶酪(cheddar)、马斯卡彭奶酪(mascarpone)、帕斯塔费拉塔奶酪(pasta filata)、马苏里拉酪(mozzarella)、波萝伏洛奶酪(provolone)、白卤奶酪(white brine cheese)、比萨奶酪(pizza cheese)、羊乳酪、布里奶酪(brie)、卡门伯特奶酪(camembert)、农家奶酪(cottage cheese)、埃丹奶酪(Edam)、高德奶酪(Gouda)、提尔西特奶酪(Tilsiter)、哈瓦蒂奶酪(Havarti)或大孔奶酪(Emmental)、瑞士奶酪(Swiss cheese)和马斯达姆奶酪(Maasdamer)。
术语“酸奶”具有其通常的含义，并且一般根据本领域众所周知的相关官方法规和标准来定义。用于制作酸奶的起子培养物包含至少一种德氏乳杆菌保加利亚亚种菌株和至少一种嗜热链球菌菌株。有趣的是，锰转运蛋白不存在于德氏乳杆菌保加利亚亚种中并且在嗜热链球菌中仅展示出低表达，这两种菌株见于酸奶的起子培养物中，使得它们对于真菌腐败特别敏感。因此，优选包括一种或多种其他细菌菌株来清除存在于酸奶中的游离锰。
在食品加工期间，化学防腐剂传统上被用来避免真菌腐败。然而，鉴于社会对较少加工且不含防腐剂的食物产品的强烈需求，本发明有助于提供一种有效的解决方案，以通过使用用于发酵食物产品和用于抑制或延迟所述食物产品中真菌生长的组合物来管理酵母和霉菌生长，所述组合物包含至多600ppm的锰和菌落形成单位/g细胞为至少2E+10的乳酸菌。
当现在公开组合物时，考虑到本公开中提供的实施例以及食物产品的性质，诸如水分活性、营养素、天然存在的锰的水平、保存期、储存条件、包装等，技术人员能够调整各种参数，诸如pH、温度和所述组合物的量以实现期望的结果。
现在所公开的组合物可在给定产品的发酵之前、开始或发酵期间添加，并且所述产品可经过进一步包装以进一步限制与酵母和霉菌的接触，和/或它还可在低温(低于15℃)储存以帮助延长保存期。
现在所公开的组合物可在给定发酵乳制品的发酵之前、开始或发酵期间添加。为了制成发酵乳制品，食品基质是乳基质。“乳基质”在本申请中被广泛地用于指基于可用作用于起子培养物的生长和发酵的培养基的乳或乳组分的组合物。“乳”通常是指通过给任何哺乳动物(诸如奶牛、绵羊、山羊、水牛或骆驼)挤奶而获得的乳分泌物。乳基质可从任何生乳材料和/或加工乳材料以及复原乳粉获得。乳基质还可以基于植物，即由植物材料制备，例如豆乳。优选由来自奶牛的乳或乳组分制备的乳基质。
乳基质包括但不限于包含蛋白质的任何乳或乳样产品的溶液/悬浮液，所述乳或乳状产品是诸如全脂乳或低脂乳、脱脂乳、酪乳、复原乳粉(reconstituted milk powder)、炼乳(condensed milk)、乳粉(dried milk)。
取决于消费者的需要，乳基质还可以是降乳糖的。乳糖降低的乳可根据本领域已知的任何方法生产，包括通过乳糖酶将乳糖水解成葡萄糖和半乳糖，或者通过纳滤、电渗析、离子交换色谱和离心。
为了使乳基质发酵，添加食品级微生物。
在添加起子培养物(现在所公开的组合物)并使乳基质经受合适的条件后，发酵工艺开始并持续一段时间。本领域普通技术人员知道如何选择合适的工艺条件，诸如温度、氧、碳水化合物的添加、一种或多种微生物的量和特征以及所需的工艺时间。这一过程可能需要三、四、五、六个小时或更长时间。
这些条件包括适合特定起子培养物菌株的温度的设定。例如，当起子培养物包含嗜中温乳酸菌时，可将温度设定为约30℃，并且如果培养物包含嗜热乳酸菌株，则将温度保持在约35℃到50℃(诸如40℃到45℃)范围内。发酵温度的设定还取决于一种或多种添加到发酵中的酶，这可由本领域普通技术人员容易地确定。在本发明的特定实施方案中，发酵温度为35℃到45℃，优选37℃到43℃，且更优选40℃到43℃。在另一实施方案中，发酵温度为15℃到35℃，优选20℃到35℃，且更优选30℃到35℃。
可使用本领域已知的任何方法终止发酵。一般来说，取决于工艺的各种参数，可通过使乳基质不适合起子培养物的一种或多种菌株生长来终止发酵。例如，当达到目标pH值时，可通过快速冷却发酵乳产品来进行终止。已知在发酵期间会发生酸化，导致形成由酪蛋白簇和链组成的三维网络。术语“目标pH值”意指发酵步骤结束时的pH值。目标pH值取决于待获得的发酵乳产品并且可由本领域普通技术人员容易地确定。
在本发明的特定实施方案中，进行发酵直到至少达到5.2的pH值，诸如直到达到5.1、5.0、4.9、4.8、4.7、4.6、4.5、4.4、4.3、4.2、4.1、4.0、3.9、3.8或3.7的pH值。优选地，进行发酵直到达到4.0到5.0、更优选4.0到4.6的目标pH值。在优选的实施方案中，进行发酵直到达到低于4.6的目标pH值。
在优选的实施方案中，发酵食物产品选自由以下组成的组：夸克、奶油奶酪、清爽奶酪、希腊酸乳、冰岛酸奶、黎巴嫩浓缩酸奶、酪乳、酸奶油、酸乳、培养乳、开菲尔(kefir)、印度酸奶、爱兰、特瓦罗格(twarog)、杜格酸奶、斯美塔那、养乐多(yakult)和达希(dahi)。
在另一优选的实施方案中，发酵食物产品是奶酪，包括欧式奶酪、新鲜奶酪、软奶酪、切达奶酪、马斯卡彭奶酪、帕斯塔费拉塔奶酪、马苏里拉奶酪、波萝伏洛奶酪、白卤奶酪、比萨奶酪、羊乳酪、布里奶酪、卡门伯特奶酪、农家奶酪、埃丹奶酪、高德奶酪、提尔西特奶酪、哈瓦蒂奶酪或大孔奶酪、瑞士奶酪和马斯达姆奶酪。
本发明的其他特征和优点将由结合附图阅读以下描述而变得显而易见。除非另有定义，否则本文所用的所有术语均具有与本领域普通技术人员通常所理解的含义相同的含义。除非本文另外指明或者上下文明显矛盾，否则在描述本发明的上下文中(尤其在下文权利要求书的上下文中)所用术语“一(a和an)”和“所述”以及类似指示物应当理解为涵盖单数与复数二者。除非另有说明，否则术语“包含”、“具有”、“包括”和“含有”应当理解为开放性术语(即，意指“包括但不限于”)。除非本文另外指明，否则本文叙述的值的范围仅仅意在作为个别地表示此范围内的每一单独值的简化方法，并且每一单独值都被并入本说明书中，就如同其在本文中被个别地叙述一样。除非本文另有指明或上下文明显矛盾，否则，本文所描述的所有方法都可按任何合适的顺序进行。除非另有说明，否则本文所提供的所有确切值均代表对应的近似值(例如，关于特定因素或测量所提供的所有确切示例性值可视为也提供对应的近似测量，在适当情况下用“约”修饰)。除非另外要求，否则，使用本文所提供的任何和所有实例或示例性用语(例如，“诸如”)仅仅意在用于更好地阐明本发明，而并不对本发明范围构成限制。本说明书中的任何用语都不应当理解为表明任何非要求保护的元素对于实施本发明是必不可少的。
实施例
本文所描述和要求保护的本发明的范围不受本文所公开的特定方面限制，因为这些方面旨在作为本发明的若干方面的说明。任何等同的方面均意在属于本发明的范围内。实际上，除了本文所示和所述的那些之外的本发明的各种修改对于本领域技术人员来说，根据前面的描述和以下的实施例将变得显而易见。此类修改也意在落入所附权利要求的范围内。在出现分歧的情况下，以本公开(包括定义)为准。
锰的定量
对于本文所公开的每种组合物，特别是在进行微波辅助消化之后，使用电感耦合等离子体质谱(ICP-MS)确定锰浓度。生长后收集细胞培养物并呈送给标准程序以获得F-DVS形式或FD-DVS形式。此外，还确定了CFU/g。本领域技术人员知道如何进行电感耦合等离子体质谱以及如何确定CFU/g。
所获得的结果示于表4中。
表4.
Figure BDA0003604753450000351
Figure BDA0003604753450000361
发酵乳样品的制备
用于本公开中的发酵乳样品如下制备。将均质化乳、特别是降脂(1.5％w/v)的均质化乳在90±1℃热处理20min并立即冷却。先前确定已存在于均质化乳中的锰浓度为约0.03ppm。将商业起子培养物(嗜热链球菌和德氏乳杆菌保加利亚亚种)以0.02％(v/w)接种在3L桶或200ml瓶中。用包含总浓度为1E+7 CFU/g的鼠李糖乳杆菌菌株(菌株1)和约30ppm的锰的组合物接种第一桶；用包含总浓度为1E+7 CFU/g的鼠李糖乳杆菌菌株(菌株1)和约195ppm的锰的组合物接种第二桶；用包含总浓度为1E+7 CFU/g的鼠李糖乳杆菌菌株(菌株1)和约625ppm的锰的组合物接种第三桶；用包含总浓度为1E+7 CFU/g的鼠李糖乳杆菌菌株(菌株3)和副干酪乳杆菌菌株以及约275ppm的锰的组合物接种第四桶；并且将第四桶用作参考且仅接种起子培养物。将所有桶在水浴中于43±1℃孵育，并且在这些条件下发酵直到达到4.60±0.1的pH。将发酵乳产品分配到200mL瓶中并冷却。
将发酵乳样品的所有部分升温到40℃的温度，并添加40ml已融化并冷却到60℃的5％无菌琼脂溶液。然后将发酵乳和琼脂的该溶液倾倒到无菌培养皿中，并将平板在LAF台中干燥30min。将这些平板用于使用霉菌的挑战(challenge)测试和使用酵母的挑战测试，其中通过评分来评价生长。
使用霉菌的挑战测试
在本公开中，使用霉菌的挑战测试如下进行。以各500个孢子/点的浓度添加不同的目标污染物(皮壳青霉、娄地青霉和潘氏青霉或卡尼青霉)。将平板在所选温度和时间孵育，并定期检查霉菌的生长。
使用酵母的挑战测试
在本公开中，使用酵母的挑战测试如下进行。通过在发酵乳样品(诸如酸奶)中接种约50CFU/g的每种目标污染物来测试不同目标污染物(德氏有孢圆酵母、汉逊德巴利酵母、长莓隐球酵母、解脂耶氏酵母)的生长或生长评分。
实施例1用包含乳酸菌和不同锰浓度的组合物抑制发酵乳产品中的霉菌
本实施例证实锰对针对不同霉菌的抑制作用的影响。使用类似于发酵乳产品的制造工艺和生产的琼脂测定法。使用鼠李糖乳杆菌和副干酪乳杆菌菌株。
图1显示3种不同霉菌(皮壳青霉、卡尼青霉和娄地青霉)在由用起子培养物(参考)或另外用包含乳酸菌和不同锰浓度的组合物(即包含乳酸菌和约30ppm的锰、或约195ppm的锰、或约275ppm的锰、或约625ppm的锰的组合物)发酵的乳制备的平板上的生长。以500个孢子/斑点的浓度添加三种目标污染物。将平板在22±1℃孵育7天。
图1证实当使用包含乳酸菌和低浓度锰的组合物时，即当使用约30ppm、约195ppm或约275ppm的锰时，对受试霉菌的抑制更显著。包含乳酸菌和约625ppm的锰的组合物仍能抑制皮壳青霉(X)和卡尼青霉(Y)的生长，但不能抑制娄地青霉(Z)的生长。此外，当不使用此类组合物时，受试霉菌会增殖，导致发酵乳产品的腐败。
图7显示3种不同霉菌在由用起子培养物(参考)或另外用包含乳酸菌的冷冻干燥(FD-DVS)DVS组合物发酵的乳制备的平板上的生长，其中FD-DVS形式具有约200ppm的锰并且所用的冷冻保护剂进一步补充有不同浓度的锰(1、5、10、20和40ppm)。向酸奶掺入皮壳青霉、卡尼青霉和娄地青霉(500个孢子/每种)并在22℃储存12天。
图7证实与锰丰富时的条件相比，在呈送给锰贫乏的条件时，受试霉菌的生长受损。因此，当将锰添加到冷冻保护剂中时，它会促进受试霉菌的生长，导致食物的腐败。因此，图7令人惊讶地显示需要具有缺失锰、尤其是缺失在其组合物中具有锰的冷冻保护剂(诸如脱脂乳粉)的现在所公开的组合物的FD-DVS形式。
总之，所有受试霉菌都在由仅用起子培养物(参考)发酵的乳制成的琼脂平板上生长得很好。因此，当不使用包含乳酸菌和锰的组合物时，受试霉菌可增殖，导致发酵乳产品的腐败。此外，图1证实，当将发酵乳产品呈送给包含乳酸菌和不同浓度的锰的组合物时，对不同受试霉菌的抑制是可能的，当使用较低浓度的锰时，效应更显著。因此，使用包含乳酸菌和低水平锰(诸如低于600ppm的锰，更优选低于275ppm，甚至更优选低于200ppm，诸如约30到约200ppm或约45ppm到约200ppm)的组合物避免发酵乳产品的腐败。
实施例2与现有技术相比，用包含乳酸菌和不同锰浓度的组合物抑制发酵乳产品 中的霉菌
还进行了本发明的组合物与现有技术之间的比较。
图5显示3种不同霉菌(皮壳青霉、潘氏青霉和娄地青霉)在由用单独的起子培养物(参考)或另外用包含与低水平的锰(诸如45或65ppm的锰)组合的乳酸菌(菌株2或1+2)的组合物或另外用基准组合物(A、B或C)发酵的乳制备的平板上的生长。基准组合物A是具有845ppm的锰的
Figure BDA0003604753450000381
XPM；基准组合物B是具有630ppm的锰的
Figure BDA0003604753450000382
YM-B Plus；基准组合物C是具有870ppm的锰的
Figure BDA0003604753450000383
YM-C。
图5证实包含乳酸菌和低浓度锰(诸如浓度低于约600ppm的锰，优选浓度为约40-600ppm的锰或浓度为约45-600ppm的锰，更优选浓度为40-70ppm的锰)的组合物是避免食物的腐败的原因，而与用于储存发酵乳样品的条件(诸如7±1℃保持24天相对于25±1℃保持11天)无关。
实施例3用包含乳酸菌和不同锰浓度的组合物抑制发酵乳产品中的酵母
本实施例证实不同酵母(诸如有孢圆酵母属或德巴利酵母属)在接种于用单独的起子培养物(参考)或另外用包含乳酸菌(诸如鼠李糖乳杆菌菌株1、鼠李糖乳杆菌菌株2、鼠李糖乳杆菌菌株3、副干酪乳杆菌)和约30ppm的锰、或约195ppm的锰、或约275ppm的锰、或约625ppm的锰的组合物发酵的发酵乳产品中时所面临的生长挑战。将生长挑战在7±1℃保持23天(图2-3)或27天(图6)。
图2和图3显示不同酵母(有孢圆酵母属和德巴利酵母属)在接种于用起子培养物、另外用包含乳酸菌和约30ppm的锰、或约195ppm的锰、或约625ppm的锰的组合物发酵的发酵乳产品中时的生长受损。对于包含约30ppm的锰的组合物，这种损害更为剧烈且持久(图2-3)。然而，含有约195ppm的锰或约625ppm的锰的组合物仍然导致受试酵母的生长受损，但程度较轻，表明包含乳酸菌和较低浓度锰的组合物确实影响真菌的生长并因此影响食物的腐败。
图6另外显示，包含乳酸菌和约275ppm的锰的组合物导致诸如德巴利酵母属的酵母的生长显著减少。所用冷冻保护剂的组成为：酪蛋白酸钠、肌醇、谷氨酸单钠、抗坏血酸钠和水，优选条件是不包括锰。所有值均以％w(成分)/w(冷冻保护剂溶液)给出。例如：酪蛋白酸钠5.55、肌醇3.75、谷氨酸单钠3.75、抗坏血酸钠5.65和水81.3，优选其中不包括锰。所有值均以％w(成分)/w(冷冻保护剂溶液)给出。
图8进一步显示德巴利酵母属在仅用起子培养物(参考)或另外用包含乳酸菌(诸如鼠李糖乳杆菌菌株2)的冷冻干燥DVS组合物接种的乳发酵产品上的生长，其中FD-DVS形式具有约200ppm的锰并且进一步补充有不同浓度的锰(1和40ppm)。因此，图8令人惊讶地显示需要具有缺失锰、尤其是缺失在其组合物中具有锰的冷冻保护剂(诸如脱脂乳粉)的组合物的FD-DVS形式。
实施例4与现有技术相比，用包含乳酸菌和不同锰浓度的组合物抑制发酵乳产品 中的酵母
还进行了本文所公开的组合物与现有技术之间的比较。
图4显示德巴利酵母属在发酵乳产品中的生长，此时所述产品仅具有单独的起子培养物(参考)，或另外具有包含与低水平的锰(诸如约45或约65ppm的锰)组合的乳酸菌(鼠李糖乳杆菌菌株2或鼠李糖乳杆菌菌株1+2)的组合物，或另外具有基准组合物(A、B或C)。基准组合物A是具有843ppm的锰的
Figure BDA0003604753450000401
XPM；基准组合物B是具有630ppm的锰的
Figure BDA0003604753450000402
YM-B Plus；基准组合物C是具有870ppm的锰的
Figure BDA0003604753450000403
YM-C。
图4证实，与包含超过600ppm的锰的基准组合物相比，当呈送给包含乳酸菌和低浓度锰(诸如45ppm或65ppm)的组合物时，德巴利酵母属在发酵乳产品上的生长显著受损。
序列表
<110> 科·汉森有限公司
<120> 用于控制真菌腐败的细菌组合物和其用途
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210 215 220
Ile Met Gly Ala Thr Ile Met Pro His Asn Ile Tyr Leu His Ser Ser
225 230 235 240
Leu Ala Gln Ser Arg Lys Tyr Asp Glu His Ile Pro Ala Gln Val Thr
245 250 255
Glu Ala Leu Arg Phe Gly Lys Trp Asp Ser Asn Val His Leu Val Ala
260 265 270
Ala Phe Leu Ile Asn Ala Leu Leu Leu Ile Leu Gly Ala Ala Leu Phe
275 280 285
Tyr Gly Val Gly Gly His Val Thr Ala Phe Gln Gly Ala Tyr Asn Gly
290 295 300
Leu Lys Asn Pro Met Ile Val Gly Gly Leu Ala Ser Pro Leu Met Ser
305 310 315 320
Thr Leu Phe Ala Phe Ala Leu Leu Ile Thr Gly Leu Ile Ser Ser Ile
325 330 335
Ala Ser Thr Leu Ala Gly Gln Ile Val Met Glu Gly Tyr Leu Asn Ile
340 345 350
Arg Met Pro Leu Trp Glu Arg Arg Leu Leu Thr Arg Leu Val Thr Leu
355 360 365
Ile Pro Ile Met Val Ile Gly Phe Met Ile Gly Phe Ser Glu His Asn
370 375 380
Phe Glu Gln Val Ile Val Tyr Ala Gln Val Ser Leu Ser Ile Ala Leu
385 390 395 400
Pro Phe Thr Leu Phe Pro Leu Val Ala Leu Thr Asn Arg Arg Asp Leu
405 410 415
Met Gly Ile His Val Asn Ser Gln Leu Val Arg Trp Val Gly Tyr Phe
420 425 430
Leu Thr Gly Val Ile Thr Val Leu Asn Ile Gln Leu Ala Ile Ser Val
435 440 445
Phe Val
450
<210> 3
<211> 535
<212> PRT
<213> 乳杆菌属
<400> 3
Met Ser Asp Asp His Lys Lys Arg His Pro Ile Lys Leu Ile Gln Tyr
1 5 10 15
Ala Asn Gly Pro Ser Leu Glu Glu Ile Asn Gly Thr Val Glu Val Pro
20 25 30
His Gly Lys Gly Phe Trp Arg Thr Leu Phe Ala Tyr Ser Gly Pro Gly
35 40 45
Ala Leu Val Ala Val Gly Tyr Met Asp Pro Gly Asn Trp Ser Thr Ser
50 55 60
Ile Thr Gly Gly Gln Asn Phe Gln Tyr Leu Leu Ile Ser Val Ile Leu
65 70 75 80
Met Ser Ser Leu Ile Ala Met Leu Leu Gln Tyr Met Ala Ala Lys Leu
85 90 95
Gly Ile Val Ser Gln Met Asp Leu Ala Gln Ala Ile Arg Ala Arg Thr
100 105 110
Ser Lys Lys Leu Gly Ile Val Leu Trp Ile Leu Thr Glu Leu Ala Ile
115 120 125
Met Ala Thr Asp Ile Ala Glu Val Ile Gly Ala Ala Ile Ala Leu Tyr
130 135 140
Leu Leu Phe His Ile Pro Leu Val Ile Ala Val Leu Val Thr Val Leu
145 150 155 160
Asp Val Leu Val Leu Leu Leu Leu Thr Lys Ile Gly Phe Arg Lys Ile
165 170 175
Glu Ala Ile Val Val Ala Leu Ile Leu Val Ile Leu Leu Val Phe Val
180 185 190
Tyr Gln Val Ala Leu Ser Asp Pro Asn Met Gly Ala Leu Leu Lys Gly
195 200 205
Phe Ile Pro Thr Gly Glu Thr Phe Ala Ser Ser Pro Ser Ile Asn Gly
210 215 220
Met Ser Pro Ile Gln Gly Ala Leu Gly Ile Ile Gly Ala Thr Val Met
225 230 235 240
Pro His Asn Leu Tyr Leu His Ser Ala Ile Ser Gln Thr Arg Lys Ile
245 250 255
Asp Tyr Lys Asn Pro Asp Asp Val Ala Gln Ala Val Lys Phe Ser Ala
260 265 270
Trp Asp Ser Asn Ile Gln Leu Ser Phe Ala Phe Val Val Asn Cys Leu
275 280 285
Leu Leu Val Met Gly Val Ala Val Phe Lys Ser Gly Ala Val Lys Asp
290 295 300
Pro Ser Phe Phe Gly Leu Phe Gln Ala Leu Ser Asp Ser Ser Thr Leu
305 310 315 320
Ser Asn Gly Val Leu Ile Ala Val Ala Lys Ser Gly Ile Leu Ser Ile
325 330 335
Leu Phe Ala Val Ala Leu Leu Ala Ser Gly Gln Asn Ser Thr Ile Thr
340 345 350
Gly Thr Leu Thr Gly Gln Val Ile Met Glu Gly Phe Val His Met Lys
355 360 365
Met Pro Leu Trp Ala Arg Arg Leu Val Thr Arg Ile Ile Ser Val Ile
370 375 380
Pro Val Ile Val Cys Val Met Leu Thr Ala Arg Asp Thr Pro Ile Gln
385 390 395 400
Gln His Glu Ala Leu Asn Thr Leu Met Asn Asn Ser Gln Val Phe Leu
405 410 415
Ala Phe Ala Leu Pro Phe Ser Met Leu Pro Leu Leu Met Phe Thr Asn
420 425 430
Ser Lys Val Glu Met Gly Asp Arg Phe Lys Asn Thr Gly Trp Val Lys
435 440 445
Val Leu Gly Trp Ile Ser Val Leu Gly Leu Thr Gly Leu Asn Leu Lys
450 455 460
Gly Leu Pro Asp Ser Ile Ala Gly Phe Phe Gly Asp His Pro Thr Ala
465 470 475 480
Thr Gln Thr Asn Met Ala Asn Ile Ile Ala Ile Val Leu Ile Val Ala
485 490 495
Ile Leu Ala Leu Leu Ala Trp Thr Ile Trp Asp Leu Tyr Lys Gly Asn
500 505 510
Gln Arg Tyr Glu Ala His Leu Ala Ala Val Ala Asp Glu Lys Glu Ala
515 520 525
Lys Ala Asp Val Asp Glu Gln
530 535
<210> 4
<211> 458
<212> PRT
<213> 干酪乳杆菌（Lactobacillus casei）
<400> 4
Met Ala Ser Glu Asp Lys Lys Ser Lys Arg Glu His Ile Ile His Phe
1 5 10 15
Glu Asp Thr Pro Ser Lys Ser Leu Asp Glu Val Asn Gly Ser Val Glu
20 25 30
Val Pro His Asn Ala Gly Phe Trp Lys Thr Leu Ala Ala Tyr Thr Val
35 40 45
Pro Gly Ile Leu Val Ala Val Gly Tyr Met Asp Pro Gly Asn Trp Ile
50 55 60
Thr Ser Ile Ala Gly Gly Ala Ser Phe Lys Tyr Ser Leu Leu Ser Val
65 70 75 80
Ile Leu Ile Ser Ser Leu Ile Ala Met Leu Leu Gln Ala Met Ala Ala
85 90 95
Arg Leu Gly Ile Val Thr Gly Arg Asp Leu Ala Gln Met Thr Arg Asp
100 105 110
His Thr Ser Lys Ala Met Gly Gly Phe Leu Trp Val Ile Thr Glu Leu
115 120 125
Ala Ile Met Ala Thr Asp Ile Ala Glu Ile Ile Gly Ser Ala Ile Ala
130 135 140
Leu Lys Leu Leu Phe Asn Met Pro Leu Ile Val Gly Ile Ile Ile Thr
145 150 155 160
Thr Ala Asp Val Leu Ile Leu Leu Leu Leu Met Arg Leu Gly Phe Arg
165 170 175
Lys Ile Glu Ala Val Val Ala Thr Leu Val Leu Val Ile Leu Leu Val
180 185 190
Phe Ala Tyr Glu Val Ile Leu Ala Gln Pro Asn Val Pro Glu Leu Leu
195 200 205
Lys Gly Tyr Leu Pro His Ala Asp Ile Val Thr Asn Lys Ser Met Leu
210 215 220
Tyr Leu Ser Leu Gly Ile Val Gly Ala Thr Val Met Pro His Asp Leu
225 230 235 240
Phe Leu Gly Ser Ser Ile Ser Gln Thr Arg Lys Ile Asp Arg Thr Lys
245 250 255
His Glu Glu Val Lys Lys Ala Ile Lys Phe Ser Thr Ile Asp Ser Asn
260 265 270
Leu Gln Leu Thr Met Ala Phe Ile Val Asn Ser Leu Leu Leu Ile Leu
275 280 285
Gly Ala Ala Leu Phe Phe Gly Thr Ser Ser Ser Val Gly Arg Phe Val
290 295 300
Asp Leu Phe Asn Ala Leu Ser Asn Ser Gln Ile Val Gly Ala Ile Ala
305 310 315 320
Ser Pro Met Leu Ser Met Leu Phe Ala Val Ala Leu Leu Ala Ser Gly
325 330 335
Gln Ser Ser Thr Ile Thr Gly Thr Leu Ala Gly Gln Ile Ile Met Glu
340 345 350
Gly Phe Ile His Leu Lys Met Pro Leu Trp Ala Gln Arg Leu Leu Thr
355 360 365
Arg Leu Met Ser Val Thr Pro Val Leu Ile Phe Ala Ile Tyr Tyr His
370 375 380
Gly Asn Glu Ala Lys Ile Glu Asn Leu Leu Thr Phe Ser Gln Val Phe
385 390 395 400
Leu Ser Ile Ala Leu Pro Phe Ala Val Ile Pro Leu Val Leu Tyr Thr
405 410 415
Ser Asp Lys Lys Ile Met Gly Glu Phe Ala Asn Arg Ala Trp Val Lys
420 425 430
Trp Thr Ala Trp Phe Ile Ser Gly Val Leu Ile Ile Leu Asn Leu Tyr
435 440 445
Leu Ile Ala Gln Thr Leu Gly Phe Val Lys
450 455
<210> 5
<211> 459
<212> PRT
<213> 短乳杆菌（Lactobacillus brevis）
<400> 5
Met Lys Asn His Glu Thr Asp Thr Lys Thr Lys His His Met Ile Glu
1 5 10 15
Ser Thr Gly Ser Gly Gln Lys Ser Leu Asp Glu Val Asn Gly Thr Val
20 25 30
Glu Val Pro Gln Asn Ala Gly Phe Trp Arg Thr Leu Met Ala Tyr Thr
35 40 45
Gly Pro Gly Ala Leu Ile Ala Val Gly Tyr Met Asp Pro Gly Asn Trp
50 55 60
Ile Thr Ser Ile Ala Gly Gly Ala Gln Tyr Lys Tyr Thr Leu Leu Thr
65 70 75 80
Val Val Leu Leu Ser Ser Leu Val Ala Met Leu Leu Gln Ala Met Ser
85 90 95
Ala Arg Leu Gly Ile Val Thr Gly Lys Asp Leu Ala Gln Leu Thr Arg
100 105 110
Glu His Thr Gly Lys Arg Thr Gly Phe Ala Leu Trp Ile Ile Thr Glu
115 120 125
Leu Ala Ile Met Ala Thr Asp Ile Ala Glu Ile Ile Gly Ser Ala Ile
130 135 140
Ala Leu Lys Leu Leu Phe Gly Phe Pro Leu Ile Val Gly Ile Ile Ile
145 150 155 160
Thr Ala Met Asp Val Leu Val Leu Leu Val Leu Met Lys Leu Gly Phe
165 170 175
Arg Lys Ile Glu Ala Ile Val Ala Thr Leu Val Ala Val Ile Leu Phe
180 185 190
Val Phe Leu Tyr Glu Val Ile Leu Ala Gln Pro His Met Gly Glu Val
195 200 205
Leu Lys Gly Tyr Leu Pro Ser Ser Thr Val Val Thr Asn His Gly Met
210 215 220
Leu Tyr Leu Ser Leu Gly Ile Val Gly Ala Thr Val Met Pro His Asp
225 230 235 240
Leu Tyr Leu Gly Ser Ser Ile Ser Gln Thr Arg Ser Phe Asp Arg Lys
245 250 255
Asn Arg Lys Ser Val Ala Gln Ala Ile Lys Phe Thr Thr Ile Asp Ser
260 265 270
Asn Ile Gln Leu Thr Leu Ala Phe Val Val Asn Ser Leu Leu Leu Ile
275 280 285
Leu Gly Ala Ala Leu Phe Phe Gly Thr Asn Ser Asp Leu Gly Arg Phe
290 295 300
Val Asp Leu Phe Asn Ala Leu Ser Asp Ser Gln Ile Val Gly Ala Ile
305 310 315 320
Ala Ser Pro Met Leu Ser Met Leu Phe Ala Leu Ala Leu Leu Ser Ser
325 330 335
Gly Gln Ser Ser Thr Ile Thr Gly Thr Leu Ala Gly Gln Ile Ile Met
340 345 350
Glu Gly Phe Ile Asn Leu Lys Met Pro Leu Trp Ala Gln Arg Leu Ile
355 360 365
Thr Arg Leu Leu Ser Val Thr Pro Val Ile Ile Phe Ala Ile Ile Tyr
370 375 380
His Gly Asn Glu Ala Lys Ile Glu Asp Leu Leu Thr Phe Ser Gln Val
385 390 395 400
Phe Leu Ser Ile Ala Leu Pro Phe Ala Met Ile Pro Leu Val Ile Phe
405 410 415
Thr Ser Ser Lys Lys Leu Met Gly Glu Phe Ala Asn Arg Thr Trp Ser
420 425 430
Lys Ile Leu Gly Trp Ile Ile Ala Val Ile Leu Ile Ile Leu Asn Ile
435 440 445
Tyr Leu Ile Leu Asn Thr Leu His Ile Val Gln
450 455
<210> 6
<211> 447
<212> PRT
<213> 乳酸片球菌（Pediococcus acidilactici）
<400> 6
Met Ser Lys Lys Leu Asp Glu Val Asp Asn Lys Ser Leu Asp Glu Ile
1 5 10 15
Asn Gly Ser Ile Lys Val Pro Lys Asn Ala Gly Phe Phe Lys Thr Leu
20 25 30
Met Ala Tyr Thr Gly Pro Gly Ile Leu Ile Ala Val Gly Tyr Met Asp
35 40 45
Pro Gly Asn Trp Ile Thr Ser Ile Ala Gly Gly Ala Gln Phe Lys Tyr
50 55 60
Thr Leu Leu Ser Val Val Leu Ile Ser Ser Leu Ile Ala Met Leu Leu
65 70 75 80
Gln Ala Met Ser Ala Arg Leu Gly Ile Val Thr Gly Lys Asp Leu Ala
85 90 95
Gln Leu Thr Arg Glu Arg Thr Ser Lys Arg Val Gly Phe Met Leu Trp
100 105 110
Val Val Ala Glu Leu Ala Ile Met Ala Thr Asp Ile Ala Glu Ile Ile
115 120 125
Gly Ser Gly Ile Ala Leu Glu Leu Leu Phe His Ile Pro Leu Ile Ile
130 135 140
Gly Ile Leu Ile Thr Ala Ala Asp Val Leu Ile Leu Leu Leu Leu Met
145 150 155 160
Arg Leu Gly Phe Arg Lys Ile Glu Ala Ile Val Ala Thr Leu Val Met
165 170 175
Val Ile Leu Ile Val Phe Ala Tyr Glu Val Phe Leu Ser Asp Pro Ser
180 185 190
Ile Ser Gly Ile Ile Lys Gly Tyr Val Pro Ala Pro Val Ile Leu Gln
195 200 205
Asn Asn Ser Met Leu Tyr Leu Ser Leu Gly Ile Val Gly Ala Thr Val
210 215 220
Met Pro His Asp Leu Tyr Leu Gly Ser Ser Ile Ser Gln Thr Arg Glu
225 230 235 240
Ile Asp Arg Arg Asp Arg Lys Asn Val Ala Gln Ala Ile Arg Phe Ser
245 250 255
Thr Ile Asp Ser Asn Met Gln Leu Phe Leu Ala Phe Ile Val Asn Ser
260 265 270
Leu Leu Leu Ile Leu Gly Ala Ala Leu Phe Tyr Gly Thr Asp Ser Ser
275 280 285
Leu Gly Arg Phe Val Asp Leu Phe Asn Ala Leu Ser Asp Asn Gln Ile
290 295 300
Val Gly Ala Ile Ala Ser Pro Met Leu Ser Met Leu Phe Ala Val Ala
305 310 315 320
Leu Leu Ala Ser Gly Gln Ser Ser Thr Ile Thr Gly Thr Leu Ser Gly
325 330 335
Gln Ile Ile Met Glu Gly Phe Ile Arg Leu Arg Val Pro Leu Trp Val
340 345 350
Gln Arg Leu Val Thr Arg Leu Leu Ser Val Ala Pro Val Leu Ile Phe
355 360 365
Ala Ile Tyr Tyr His Gly Asp Glu Ala Lys Ile Glu Asn Leu Leu Thr
370 375 380
Phe Ser Gln Val Phe Leu Ser Val Ala Leu Pro Phe Ala Val Ile Pro
385 390 395 400
Leu Val Met Tyr Thr Ser Ser Lys Lys Leu Met Gly Glu Phe Ala Asn
405 410 415
Arg Gln Trp Val Lys Trp Cys Ala Trp Ile Ala Thr Ile Ile Leu Ile
420 425 430
Leu Leu Asn Ile Tyr Leu Ile Leu Gln Thr Leu Gly Ile Val Lys
435 440 445
<210> 7
<211> 464
<212> PRT
<213> 植物乳杆菌（Lactobacillus plantarum）
<400> 7
Met Lys Ser Ala Lys Thr Lys Asp His Ala Lys Met Lys Ala Ala Glu
1 5 10 15
Glu Lys Ala Ile His Ser Thr Gly Ala Asp Ser Lys Ser Leu Asp Glu
20 25 30
Val Asn Gly Ser Val Arg Val Pro Lys Asp Ala Ser Phe Trp Arg Thr
35 40 45
Leu Ile Ala Tyr Thr Gly Pro Gly Ala Leu Val Ala Val Gly Tyr Met
50 55 60
Asp Pro Gly Asn Trp Ile Thr Ser Ile Ala Gly Gly Ser Gln Tyr Lys
65 70 75 80
Tyr Ala Leu Leu Ser Val Ile Leu Leu Ser Ser Leu Ile Ala Met Leu
85 90 95
Leu Gln Ala Met Ala Ala Arg Leu Gly Ile Val Thr Gly Lys Asp Leu
100 105 110
Ala Gln Leu Thr Arg Glu Arg Thr Ser Lys Gly Met Gly Ile Phe Leu
115 120 125
Trp Ile Ile Thr Glu Leu Ala Ile Met Ala Thr Asp Val Ala Glu Ile
130 135 140
Ile Gly Ser Gly Ile Ala Leu Lys Leu Leu Phe Gly Phe Pro Leu Ile
145 150 155 160
Val Gly Ile Leu Ile Thr Thr Ala Asp Val Leu Ile Leu Leu Leu Leu
165 170 175
Met Lys Leu Gly Phe Arg Lys Ile Glu Ala Ile Val Ala Thr Leu Val
180 185 190
Ala Val Ile Leu Phe Val Phe Leu Tyr Glu Val Ile Ile Ser Gln Pro
195 200 205
Asn Ile Pro Glu Met Leu Lys Gly Tyr Val Pro Thr Ser Arg Ile Val
210 215 220
Ser Asn Arg Ser Met Leu Phe Leu Ala Leu Gly Ile Val Gly Ala Thr
225 230 235 240
Val Met Pro His Asn Leu Tyr Leu Gly Ser Ser Ile Ser Gln Thr Arg
245 250 255
Gln Val Asp Arg Ser Asp Glu Lys Glu Val Ala Lys Ala Val Lys Phe
260 265 270
Thr Thr Ile Asp Ser Asn Ile Gln Leu Ser Val Ala Phe Val Val Asn
275 280 285
Ser Leu Leu Leu Ile Leu Gly Ala Ala Leu Phe Phe Gly Thr Lys Gly
290 295 300
Asp Leu Gly Arg Phe Val Asp Leu Tyr Asn Ala Leu Gly Asp Ser Lys
305 310 315 320
Val Val Gly Ser Ile Ala Ser Pro Leu Leu Ser Met Leu Phe Ala Ile
325 330 335
Ala Leu Leu Ser Ser Gly Gln Ser Ser Thr Ile Thr Gly Thr Leu Ser
340 345 350
Gly Gln Ile Ile Met Glu Gly Phe Ile Arg Leu Lys Met Pro Leu Trp
355 360 365
Ala Gln Arg Leu Leu Thr Arg Leu Ile Ser Val Thr Pro Val Leu Ala
370 375 380
Phe Ala Ile Tyr Tyr His Gly Asn Glu Ala Lys Ile Glu Asp Leu Leu
385 390 395 400
Thr Met Ser Gln Val Phe Leu Ser Ile Ala Leu Pro Phe Ala Met Ile
405 410 415
Pro Leu Val Met Phe Thr Ser Asn Arg Ala Leu Met Gly Asn Phe Thr
420 425 430
Asn Arg Val Trp Val Lys Trp Thr Ala Trp Ile Val Thr Val Ile Leu
435 440 445
Ile Ile Leu Asn Ile Tyr Leu Ile Leu Gln Thr Val Gly Leu Val Lys
450 455 460
<210> 8
<211> 443
<212> PRT
<213> 沙克乳杆菌（Lactobacillus sakei）
<400> 8
Met His Tyr Ala Asp Gly Ser Ser Leu Glu Glu Ile Asn Asn Thr Val
1 5 10 15
Ala Ile Pro Lys Asn Ala Gly Phe Trp Lys Thr Leu Met Ala Phe Met
20 25 30
Gly Pro Gly Ala Leu Val Ala Val Gly Tyr Met Asp Pro Gly Asn Trp
35 40 45
Ile Thr Ser Ile Ala Gly Gly Ala Gln Phe Ala Tyr Thr Leu Ile Ser
50 55 60
Val Ile Leu Val Ser Asn Leu Ile Ala Met Leu Leu Gln Ala Met Ala
65 70 75 80
Ala Arg Leu Gly Ile Val Thr Gly Met Asp Leu Ala Gln Met Thr Arg
85 90 95
Ala Lys Thr Gly Lys Lys Met Gly Ile Phe Leu Trp Ile Val Thr Glu
100 105 110
Leu Ala Ile Met Ala Thr Asp Ile Ala Glu Ile Ile Gly Ser Ala Ile
115 120 125
Ala Leu Glu Leu Ile Phe Asn Ile Pro Leu Leu Trp Gly Val Leu Ile
130 135 140
Thr Ala Phe Asp Val Leu Leu Leu Leu Leu Leu Met Lys Leu Gly Phe
145 150 155 160
Arg Lys Ile Glu Ala Ile Val Ala Thr Leu Val Ala Val Ile Leu Phe
165 170 175
Val Phe Leu Tyr Glu Val Ile Leu Ala Gln Pro Asn Met Gly Asp Val
180 185 190
Val Arg Gly Phe Val Pro Ser Pro Arg Ile Met Thr Asp Lys Lys Met
195 200 205
Leu Phe Leu Ala Leu Gly Ile Val Gly Ala Thr Val Met Pro His Asn
210 215 220
Leu Tyr Leu His Ser Ser Ile Ala Gln Ala Arg Gln Tyr Asp Arg Asp
225 230 235 240
Asp Val Ala Glu Lys Arg Lys Ala Ile Lys Phe Thr Val Ile Asp Ser
245 250 255
Asn Ile Gln Leu Thr Ile Ala Phe Val Val Asn Cys Leu Leu Leu Ile
260 265 270
Leu Gly Ala Ala Met Phe Tyr Gly Thr Asn Ser Asp Leu Gly Arg Phe
275 280 285
Val Asp Leu Phe Asn Ala Leu Gln Asn Lys Glu Ile Val Gly Ser Ile
290 295 300
Ala Ser Pro Met Leu Ser Leu Leu Phe Ala Val Ala Leu Leu Ala Ser
305 310 315 320
Gly Gln Asn Ser Thr Ile Thr Gly Thr Leu Ser Gly Gln Ile Val Met
325 330 335
Glu Gly Phe Val Arg Met Lys Ile Pro Leu Trp Ala Arg Arg Val Ile
340 345 350
Thr Arg Gly Leu Ser Ile Leu Pro Val Ile Ile Phe Thr Val Tyr Tyr
355 360 365
His Gly Asn Glu Ala Gln Val Glu Asn Leu Leu Ile Tyr Ser Gln Val
370 375 380
Phe Leu Ser Ile Ala Leu Pro Val Ser Met Ile Pro Leu Thr Leu Phe
385 390 395 400
Thr Ser Asp Glu Lys Ile Met Gly Pro Phe Val Asn Arg Pro Trp Val
405 410 415
Lys Tyr Thr Ala Trp Phe Val Thr Ile Val Leu Thr Leu Leu Asn Ile
420 425 430
Tyr Leu Ile Leu Gln Thr Val Gly Leu Ala Ala
435 440
<210> 9
<211> 454
<212> PRT
<213> 消化乳杆菌（Lactobacillus alimentarius）
<400> 9
Met Ser Ser Lys Asn Lys Lys His Glu Ser Leu Ile His Tyr Ala Asn
1 5 10 15
Gly Pro Ser Leu Glu Glu Ile Asn Asp Thr Val Glu Ile Pro Lys Asp
20 25 30
Ala Gly Phe Phe Lys Thr Leu Leu Ala Tyr Ser Gly Pro Gly Ala Leu
35 40 45
Val Ala Val Gly Tyr Met Asp Pro Gly Asn Trp Val Thr Ser Ile Ala
50 55 60
Gly Gly Ala Gln Phe Lys Tyr Lys Leu Leu Ser Val Ile Leu Ile Ser
65 70 75 80
Ser Leu Ile Ala Met Leu Leu Gln Tyr Met Ser Ala Lys Leu Gly Ile
85 90 95
Val Thr Gly Arg Asp Leu Ala Gln Leu Thr Arg Asp Arg Thr Ser Arg
100 105 110
Val Gly Gly Phe Ile Leu Trp Ile Ile Thr Glu Leu Ala Ile Met Ala
115 120 125
Thr Asp Ile Ala Glu Ile Ile Gly Ser Ala Ile Ala Leu Lys Leu Leu
130 135 140
Phe Asn Ile Pro Val Leu Trp Gly Val Ile Ile Thr Ala Phe Asp Val
145 150 155 160
Leu Leu Leu Leu Val Leu Met Lys Leu Gly Phe Arg Lys Ile Glu Ala
165 170 175
Ile Val Ala Thr Leu Ile Met Val Ile Leu Leu Val Phe Leu Tyr Glu
180 185 190
Val Ile Leu Ala Lys Pro Asp Val Gly Gln Met Met Val Gly Phe Ile
195 200 205
Pro Glu Pro Lys Ile Leu Gln Asn Gln Ser Met Leu Tyr Leu Ser Leu
210 215 220
Gly Ile Val Gly Ala Thr Val Met Pro His Asn Leu Tyr Leu His Ser
225 230 235 240
Ser Ile Ser Gln Ala Arg Lys Tyr Asp Arg Asp Asp Pro Lys Ser Ile
245 250 255
His Gln Ala Val Arg Phe Ser Thr Trp Asp Ser Asn Ile Gln Leu Thr
260 265 270
Leu Ala Phe Val Val Asn Thr Leu Leu Leu Leu Leu Gly Ala Ala Leu
275 280 285
Phe Tyr Gly Thr Ser Ser Asp Leu Gly Arg Phe Val Asp Leu Phe Asn
290 295 300
Ala Leu Gln Asp Pro Lys Val Ala Gly Ala Val Ala Ser Pro Val Leu
305 310 315 320
Ser Ile Leu Phe Ala Val Ala Leu Leu Ala Ser Gly Gln Asn Ser Thr
325 330 335
Ile Thr Gly Thr Leu Ser Gly Gln Ile Val Met Glu Gly Phe Ile His
340 345 350
Met Lys Met Lys Leu Trp Ala Arg Arg Val Ile Thr Arg Leu Met Ser
355 360 365
Ile Ile Pro Val Ile Thr Phe Ala Ile Ile Tyr His Gly Asn Glu Ala
370 375 380
Lys Ile Glu Ser Leu Leu Thr Phe Ser Gln Val Phe Leu Ser Val Ala
385 390 395 400
Leu Pro Phe Ser Ile Phe Pro Leu Ile Lys Phe Thr Ser Asn Lys Lys
405 410 415
Leu Met Gly Glu Phe Val Asn Asn Lys Leu Val Glu Tyr Ile Gly Tyr
420 425 430
Phe Val Ala Ile Val Leu Thr Ile Leu Asn Ile Trp Leu Ile Tyr Thr
435 440 445
Thr Phe Val Pro Thr Ala
450
<210> 10
<211> 451
<212> PRT
<213> 栖花乳杆菌（Lactobacillus floricola）
<400> 10
Met Thr Lys Glu Glu Thr Lys Leu Phe His Tyr Ala Asp Gly Pro Ser
1 5 10 15
Leu Glu Glu Ile Asn Gly Thr Val Ala Val Pro Lys Lys Gly Gly Phe
20 25 30
Trp Lys Thr Leu Phe Ala Phe Ser Gly Pro Gly Ala Leu Val Ala Val
35 40 45
Gly Tyr Met Asp Pro Gly Asn Trp Val Thr Ser Ile Ala Gly Gly Ala
50 55 60
Gln Tyr Gln Tyr Thr Leu Leu Ser Val Ile Leu Ile Ser Ser Leu Ile
65 70 75 80
Ala Met Leu Leu Gln Ala Met Ser Ala Arg Leu Gly Ile Ala Ser Gly
85 90 95
Leu Asp Leu Ala Gln Ala Thr Ala Lys His Ser Pro Lys Trp Leu Arg
100 105 110
Tyr Thr Leu Trp Ile Ile Thr Glu Leu Ala Ile Met Ala Thr Asp Ile
115 120 125
Ala Glu Ile Val Gly Ala Ala Ile Ala Leu Lys Leu Leu Phe Asn Leu
130 135 140
Pro Leu Ile Val Gly Ile Phe Leu Thr Thr Leu Asp Val Met Leu Leu
145 150 155 160
Leu Leu Leu Met Lys Leu Gly Phe Arg Lys Ile Glu Ala Ile Val Gly
165 170 175
Ala Leu Ile Val Ser Ile Leu Val Ile Phe Leu Tyr Glu Val Ile Leu
180 185 190
Ala Arg Pro Asp Val Gly Ala Met Phe Ala Gly Tyr Ile Pro Gln Pro
195 200 205
Glu Val Val Thr Asn Lys Gly Ala Phe Tyr Ile Ala Leu Gly Ile Val
210 215 220
Gly Ala Thr Val Met Pro His Asn Leu Tyr Leu His Ser Ser Ile Ala
225 230 235 240
Gln Ala Arg Gln Tyr Asp Arg Asn Asp Ile Glu Glu Lys Lys Arg Ala
245 250 255
Ile Lys Phe Thr Val Leu Asp Ser Asn Ile Gln Leu Ser Val Ala Phe
260 265 270
Val Val Asn Thr Leu Leu Leu Leu Leu Gly Ala Ala Leu Phe Tyr Gly
275 280 285
Ala Gln Thr Asp Leu Gly Thr Phe Ser Glu Leu Tyr Asn Ala Leu Gln
290 295 300
Asn Pro Gln Val Ala Gly Val Ile Ala Ser Pro Ile Leu Ser Val Leu
305 310 315 320
Phe Ala Val Ala Leu Leu Ala Ser Gly Gln Asn Ser Thr Ile Thr Gly
325 330 335
Thr Leu Ser Gly Gln Ile Val Met Glu Gly Phe Ile His Leu Lys Met
340 345 350
Pro Met Trp Ala Arg Arg Val Ile Thr Arg Leu Ile Ser Val Ile Pro
355 360 365
Val Leu Ile Phe Ala Ile Ile Tyr His Ser Asn Glu Ala Lys Ile Glu
370 375 380
Asp Leu Leu Val Phe Ser Gln Val Phe Leu Ser Ile Ala Leu Pro Val
385 390 395 400
Ser Ile Ile Pro Leu Val Met Phe Thr Ala Asn Lys Lys Ile Met Gly
405 410 415
Pro Phe Val Asn Lys Lys Trp Val Thr Ile Thr Ser Ser Leu Val Ala
420 425 430
Ile Ile Leu Thr Gly Leu Asn Ile Phe Leu Ile Leu Gln Thr Leu Gly
435 440 445
Trp Val Gln
450
<210> 11
<211> 455
<212> PRT
<213> 短乳杆菌（Lactobacillus brevis）
<400> 11
Met Thr Asp Asn Val Ser Ala Lys Ser Val Gln Gly Asp Leu Thr Asn
1 5 10 15
Gly Pro Ser Leu Ala Glu Ile Asn Gly Ser Val Arg Val Pro Lys Glu
20 25 30
Lys Gly Phe Val Arg Asn Leu Leu Ala Phe Ser Gly Pro Gly Ala Leu
35 40 45
Val Ala Val Gly Tyr Met Asp Pro Gly Asn Trp Val Thr Ser Ile Gly
50 55 60
Gly Gly Ala Gln Tyr Gly Tyr Leu Leu Met Ser Val Ile Leu Met Ser
65 70 75 80
Ser Leu Ile Ala Met Leu Leu Gln Tyr Met Ala Ala Lys Leu Gly Ile
85 90 95
Val Thr Gln Met Asp Leu Ala Arg Ala Thr Arg Ala His Thr Gly Lys
100 105 110
Arg Ile Gly Ala Val Leu Trp Val Met Thr Glu Leu Ala Ile Met Ala
115 120 125
Thr Asp Ile Ala Glu Val Ile Gly Gly Ala Ile Ala Leu Lys Leu Leu
130 135 140
Phe Gly Val Pro Leu Ile Leu Gly Val Ser Leu Thr Val Leu Asp Val
145 150 155 160
Leu Leu Leu Leu Leu Leu Thr Arg Leu Gly Phe Arg Lys Ile Glu Ala
165 170 175
Ile Val Leu Cys Leu Ile Leu Val Ile Leu Val Val Phe Ala Tyr Glu
180 185 190
Val Val Ile Ala Gln Pro Ser Met Gly Gln Ala Val Ala Ser Phe Val
195 200 205
Pro Gln Ala Glu Ile Met Arg Pro Gly Gln Leu Thr Met Ala Leu Gly
210 215 220
Ile Val Gly Ala Thr Val Met Pro His Asn Leu Tyr Leu His Ser Ser
225 230 235 240
Ile Ala Gln Thr Arg Lys Phe Asp Arg Gln Asp Pro Ala Glu Met Ala
245 250 255
Arg Ala Val Lys Phe Thr Ala Trp Asp Ser Asn Ile Gln Leu Phe Gly
260 265 270
Ala Phe Ile Ile Asn Cys Leu Leu Leu Leu Leu Gly Ala Ala Met Phe
275 280 285
Phe Gly Lys Asp Ala Gly Ala Leu Gly Thr Phe Gly Gln Leu Tyr Asp
290 295 300
Ala Leu Gln Asp Asn Arg Leu Ala Gly Ala Val Ala Ser Pro Val Leu
305 310 315 320
Ser Thr Leu Phe Ala Val Ala Leu Leu Ala Ser Gly Gln Asn Ser Thr
325 330 335
Ile Thr Gly Thr Leu Thr Gly Gln Val Ile Met Glu Gly Phe Ile Asn
340 345 350
Met Arg Leu Pro Ile Trp Val Arg Arg Leu Val Thr Arg Leu Ile Ser
355 360 365
Val Ala Pro Val Ile Ile Val Thr Ile Leu Tyr Gly Gly Ser Glu Gln
370 375 380
Ala Leu Asp Arg Leu Leu Val Asn Ser Gln Val Phe Leu Ser Ile Ala
385 390 395 400
Leu Pro Phe Ser Met Ile Pro Leu Thr Ile Phe Thr Ser Ser Lys Arg
405 410 415
Ile Met Gly Thr Arg Trp Val Asn Arg Trp Trp Val Thr Ala Leu Ala
420 425 430
Trp Gly Cys Thr Ala Ile Leu Thr Val Leu Asn Ile Gln Ile Val Trp
435 440 445
Ala Thr Met Thr Thr Leu Phe
450 455
<210> 12
<211> 450
<212> PRT
<213> 干酪乳杆菌
<400> 12
Met Ala Arg Pro Asp Glu Arg Leu Thr Val Gln Arg Glu Lys Arg Ser
1 5 10 15
Leu Asp Asp Ile Asn Arg Ser Val Gln Val Pro Ser Val Tyr Glu Ser
20 25 30
Ser Phe Phe Gln Lys Phe Leu Ala Tyr Ser Gly Pro Gly Ala Leu Val
35 40 45
Ala Val Gly Tyr Met Asp Pro Gly Asn Trp Leu Thr Ala Leu Glu Gly
50 55 60
Gly Ser Arg Tyr His Tyr Ala Leu Leu Ser Val Leu Leu Met Ser Ile
65 70 75 80
Leu Val Ala Met Phe Met Gln Thr Leu Ala Ile Lys Leu Gly Val Val
85 90 95
Ala Arg Leu Asp Leu Ala Gln Ala Ile Ala Ala Phe Ile Pro His Trp
100 105 110
Ser Arg Ile Cys Leu Trp Leu Ile Asn Glu Ala Ala Met Met Ala Thr
115 120 125
Asp Met Thr Gly Val Val Gly Thr Ala Ile Ala Leu Lys Leu Leu Phe
130 135 140
Gly Leu Pro Leu Met Trp Gly Met Leu Leu Thr Ile Ala Asp Val Leu
145 150 155 160
Val Val Leu Leu Phe Leu Arg Phe Gly Ile Arg Arg Val Glu Leu Ile
165 170 175
Val Leu Val Ser Ile Leu Thr Val Gly Ile Ile Phe Gly Ile Glu Val
180 185 190
Ala Arg Ala Asp Pro Ser Ile Gly Gly Ile Ala Gly Gly Phe Val Pro
195 200 205
His Thr Asp Ile Leu Thr Asn His Gly Met Leu Leu Leu Ser Leu Gly
210 215 220
Ile Met Gly Ala Thr Ile Met Pro His Asn Ile Tyr Leu His Ser Ser
225 230 235 240
Leu Ala Gln Ser Arg Lys Tyr Asp Glu His Ile Pro Ala Gln Val Thr
245 250 255
Glu Ala Leu Arg Phe Gly Lys Trp Asp Ser Asn Val His Leu Val Ala
260 265 270
Ala Phe Leu Ile Asn Ala Leu Leu Leu Ile Leu Gly Ala Ala Leu Phe
275 280 285
Tyr Gly Val Gly Gly His Val Thr Ala Phe Gln Gly Val Tyr Asn Gly
290 295 300
Leu Lys Asn Pro Met Ile Val Gly Gly Leu Ala Ser Pro Leu Met Ser
305 310 315 320
Thr Leu Phe Ala Phe Ala Leu Leu Ile Thr Gly Leu Ile Ser Ser Ile
325 330 335
Ala Ser Thr Leu Ala Gly Gln Ile Val Met Glu Gly Tyr Leu Asn Ile
340 345 350
Arg Met Pro Leu Trp Glu Arg Arg Leu Leu Thr Arg Leu Val Thr Leu
355 360 365
Ile Pro Ile Met Val Ile Gly Phe Met Ile Gly Phe Ser Glu His Asn
370 375 380
Phe Glu Gln Val Ile Val Tyr Ala Gln Val Ser Leu Ser Ile Ala Leu
385 390 395 400
Pro Phe Thr Leu Phe Pro Leu Val Ala Leu Thr Asn Arg Arg Asp Leu
405 410 415
Met Gly Ile His Val Asn Ser Gln Leu Val Arg Trp Val Gly Tyr Phe
420 425 430
Leu Thr Gly Val Ile Thr Val Leu Asn Ile Gln Leu Ala Ile Ser Val
435 440 445
Phe Val
450
<210> 13
<211> 451
<212> PRT
<213> 鼠李糖乳杆菌（Lactobacillus rhamnosus）
<400> 13
Met Thr Lys Arg Asn Glu Gln Leu Ser Val Gln Gln Ala Lys Pro Ser
1 5 10 15
Leu Asp Glu Ile Asn Arg Ser Val Gln Val Pro Gly Val Tyr Glu Pro
20 25 30
Ser Phe Val Gln Lys Phe Leu Ala Tyr Ser Gly Pro Gly Ala Leu Val
35 40 45
Ala Val Gly Tyr Met Asp Pro Gly Asn Trp Leu Thr Ala Leu Glu Gly
50 55 60
Gly Ser Arg Tyr His Glu Thr Leu Leu Ala Val Leu Leu Leu Ser Ile
65 70 75 80
Leu Ala Ala Met Phe Met Gln Thr Leu Ala Ile Lys Leu Gly Val Val
85 90 95
Ala Arg Leu Asp Leu Ala Gln Ala Ile Ala Ala Phe Val Pro Lys Trp
100 105 110
Ser Arg Ile Gly Leu Trp Leu Val Asn Glu Ala Ala Met Met Ala Thr
115 120 125
Asp Met Thr Gly Val Val Gly Thr Ala Ile Ala Leu Lys Leu Leu Phe
130 135 140
Gly Leu Pro Leu Met Trp Gly Met Leu Leu Thr Ile Ala Asp Val Leu
145 150 155 160
Val Val Leu Met Phe Leu Arg Phe Gly Ile Arg Arg Ile Glu Leu Ile
165 170 175
Val Leu Ala Ser Ile Leu Thr Val Gly Ile Ile Phe Gly Ile Glu Val
180 185 190
Val Arg Ala Arg Pro Ser Met Gly Gly Ile Val Ala Gly Leu Val Pro
195 200 205
His Thr Glu Ile Leu Thr Asn Arg Gly Met Leu Leu Leu Ser Leu Gly
210 215 220
Ile Met Gly Ala Thr Ile Met Pro His Asn Ile Tyr Leu His Ser Ser
225 230 235 240
Leu Ala Gln Ser Arg Arg Tyr Asp Glu His Ile Pro Ala Gln Val Thr
245 250 255
Glu Ala Leu Arg Phe Gly Lys Trp Asp Ser Asn Val His Leu Val Ala
260 265 270
Ala Phe Ile Ile Asn Ala Leu Leu Leu Ile Leu Gly Ala Thr Leu Phe
275 280 285
Tyr Gly Met Ser Ser His Ala Thr Ala Phe Glu Gly Val Tyr Asn Gly
290 295 300
Leu Lys Asn Pro Ala Ile Val Gly Gly Leu Ala Ser Pro Leu Met Ser
305 310 315 320
Thr Leu Phe Ala Phe Ala Leu Leu Ile Thr Gly Leu Ile Ser Ser Ile
325 330 335
Ala Ser Thr Leu Ala Gly Gln Ile Val Met Glu Gly Tyr Leu Asn Ile
340 345 350
Gln Ile Pro Leu Trp Ala Arg Arg Leu Leu Thr Arg Leu Val Thr Leu
355 360 365
Ile Pro Ile Met Ile Ile Gly Phe Val Met Gly Phe Ser Glu Gln His
370 375 380
Phe Glu Gln Val Ile Val Tyr Ala Gln Val Ala Leu Ser Ile Ala Leu
385 390 395 400
Pro Phe Thr Leu Phe Pro Leu Val Ala Leu Thr Asp Arg Arg Asp Leu
405 410 415
Met Gly Gln His Val Asn Ser Pro Val Val Arg Trp Met Gly Tyr Val
420 425 430
Leu Thr Gly Ile Ile Thr Leu Leu Asn Val Gln Leu Ile Leu Ser Val
435 440 445
Ile Leu Pro
450
<210> 14
<211> 442
<212> PRT
<213> 开菲尔乳杆菌（Lactobacillus kefiri）
<400> 14
Met Ser Gln Glu Pro Thr His Lys Ser Leu Asp Glu Ile Asn Gln Ser
1 5 10 15
Val Glu Val Pro Ser Val Tyr Glu Thr Ser Phe Leu Gln Lys Phe Leu
20 25 30
Ala Tyr Ser Gly Pro Gly Ala Leu Val Ala Val Gly Tyr Met Asp Pro
35 40 45
Gly Asn Trp Leu Thr Ser Leu Ser Gly Gly Ser Gln Phe Arg Tyr Ala
50 55 60
Leu Leu Ser Val Leu Leu Met Ser Ile Leu Val Ala Met Phe Met Gln
65 70 75 80
Thr Leu Ser Ile Lys Leu Gly Val Val Ala Arg Leu Asp Leu Ala Gln
85 90 95
Ala Ile Ala Gln Lys Val Pro Lys Ser Gly Arg Tyr Thr Leu Trp Ile
100 105 110
Ile Asn Glu Leu Ala Met Met Ala Thr Asp Met Thr Gly Val Val Gly
115 120 125
Thr Ala Ile Ala Leu Lys Leu Leu Phe Gly Leu Pro Leu Val Tyr Gly
130 135 140
Ile Leu Leu Thr Ile Phe Asp Val Leu Leu Val Leu Leu Phe Leu Arg
145 150 155 160
Phe Gly Ile Arg Arg Ile Glu Phe Ile Val Leu Ala Ala Ile Leu Ile
165 170 175
Val Gly Val Ile Phe Gly Ile Glu Val Thr Arg Ala Thr Pro Asn Ile
180 185 190
Val Glu Ile Ala Gly Gly Leu Ile Pro Thr Thr His Ile Val Thr Asn
195 200 205
His Glu Met Leu Ile Met Ser Leu Gly Ile Val Gly Ala Thr Ile Met
210 215 220
Pro His Asn Val Tyr Leu His Ser Ser Leu Ala Gln Ser Arg Arg Tyr
225 230 235 240
Asp Tyr His Asn Pro Lys Gln Val Asn Glu Ala Leu Arg Phe Ala Lys
245 250 255
Trp Asp Ser Asn Val His Leu Val Ala Ala Phe Leu Ile Asn Ala Leu
260 265 270
Leu Leu Val Leu Gly Gly Thr Leu Phe Phe His Thr Asn Ser His Phe
275 280 285
Ser Ala Phe Gln Asp Val Tyr Asn Gly Leu Lys Ser Ser Ala Ile Val
290 295 300
Gly Ser Leu Ala Ser Pro Leu Met Ser Thr Leu Phe Ala Phe Ala Leu
305 310 315 320
Leu Ile Thr Gly Met Ile Ser Ser Ile Thr Ser Thr Leu Ser Gly Gln
325 330 335
Ile Val Met Glu Gly Tyr Leu His Ile Arg Leu Pro Leu Trp Glu Arg
340 345 350
Arg Leu Leu Thr Arg Phe Val Thr Leu Ile Pro Ile Leu Ala Ile Gly
355 360 365
Phe Leu Val Gly Phe Asn Asp His Asp Phe Glu Glu Ile Ile Val Tyr
370 375 380
Ala Gln Ile Ala Leu Ser Ile Ala Leu Pro Phe Thr Leu Phe Pro Met
385 390 395 400
Val Ala Leu Thr Ser Asn His Asp Leu Met Gly Val His Thr Asn Arg
405 410 415
Arg Tyr Val Thr Val Ile Gly Tyr Leu Leu Thr Ser Ile Ile Thr Ile
420 425 430
Leu Asn Leu Gln Phe Val Leu Ala Ser Ile
435 440
<210> 15
<211> 439
<212> PRT
<213> 消化乳杆菌
<400> 15
Met Pro Asn Lys Lys Ser Leu Asp Glu Ile Asn Glu Ser Val Lys Val
1 5 10 15
Pro Ser Val Tyr Asp Thr Ser Phe Leu Gln Lys Phe Leu Ala Tyr Ser
20 25 30
Gly Pro Gly Ala Leu Val Ala Val Gly Tyr Met Asp Pro Gly Asn Trp
35 40 45
Leu Thr Ser Leu Ser Gly Gly Ser Gln Tyr Arg Tyr Asp Leu Leu Ser
50 55 60
Val Leu Leu Ile Ser Ile Leu Val Ala Met Phe Met Gln Thr Leu Ser
65 70 75 80
Ile Lys Leu Gly Val Val Ala Arg Leu Asp Leu Ala Gln Ala Ile Ala
85 90 95
Thr Lys Val Ser Lys Pro Ile Arg Tyr Phe Leu Trp Ile Leu Asn Glu
100 105 110
Ile Ala Met Met Ala Thr Asp Leu Thr Gly Val Ile Gly Thr Ala Ile
115 120 125
Ala Leu Lys Leu Leu Phe Asn Leu Pro Leu Val Phe Gly Ile Leu Leu
130 135 140
Thr Val Phe Asp Val Leu Ile Val Leu Ile Phe Leu Arg Phe Gly Ile
145 150 155 160
Arg Arg Ile Glu Phe Ile Val Leu Ala Ala Ile Leu Thr Val Gly Ile
165 170 175
Ile Phe Gly Ile Glu Val Phe Arg Ala Gln Pro Lys Leu Phe Ser Ile
180 185 190
Ile Ser Gly Val Ile Pro Ser Thr Asp Leu Phe Thr Asn His Arg Lys
195 200 205
Leu Val Leu Ser Leu Gly Ile Val Gly Ala Thr Ile Met Pro His Asn
210 215 220
Ile Tyr Leu His Ser Ser Leu Ala Gln Ser Arg Arg Tyr Asp His Asn
225 230 235 240
Asp Pro Leu Gln Val Asn Glu Ala Leu Arg Phe Ala Lys Trp Asp Ser
245 250 255
Asn Val His Leu Ile Ala Ala Phe Ile Ile Asn Ala Leu Leu Leu Val
260 265 270
Leu Gly Gly Thr Leu Phe Tyr His Met Thr Asn Gln Leu Ala Ser Leu
275 280 285
Gln Asp Val Phe Thr Gly Leu Lys Ser His Ala Ile Val Gly Thr Leu
290 295 300
Ala Ser Pro Leu Met Ser Trp Leu Phe Ala Phe Ala Leu Leu Ile Thr
305 310 315 320
Gly Met Ile Ser Ser Ile Thr Ser Thr Leu Ser Gly Gln Ile Val Met
325 330 335
Glu Gly Tyr Leu Asn Ile Arg Leu Pro Leu Trp Gln Arg Arg Leu Leu
340 345 350
Thr Arg Phe Val Thr Leu Ile Pro Ile Leu Ile Ile Gly Phe Ile Val
355 360 365
His Phe Asn Glu Gln Asp Phe Glu Asn Leu Ile Val Tyr Ala Gln Ile
370 375 380
Ile Leu Ser Ile Ala Leu Pro Phe Thr Leu Phe Pro Met Ile Phe Leu
385 390 395 400
Thr Asn Asp Lys Lys Ile Met Gly Asn His Val Asn Ser Lys Leu Thr
405 410 415
Thr Thr Val Gly Ile Ile Leu Ala Ser Ala Ile Thr Ile Leu Asn Leu
420 425 430
Gln Leu Leu Phe Ser Leu Ile
435
<210> 16
<211> 441
<212> PRT
<213> 植物乳杆菌
<400> 16
Met Gln Ser His Arg His Gln Ser Leu Glu Glu Ile Asn Gln Ser Val
1 5 10 15
Ala Val Pro Asp Val His Gln Thr Ala Phe Trp Arg Lys Phe Leu Ala
20 25 30
Tyr Ser Gly Pro Gly Ala Leu Val Ala Val Gly Tyr Met Asp Pro Gly
35 40 45
Asn Trp Leu Thr Ser Leu Ala Gly Gly Gly Gln Phe Gln Tyr Arg Leu
50 55 60
Leu Ala Val Leu Ala Leu Ala Ile Ile Val Ala Met Phe Met Gln Gly
65 70 75 80
Leu Ala Ile Arg Leu Gly Val Val Ala Arg Gln Asp Leu Ala Gln Ala
85 90 95
Ile Ala Ser Lys Leu Pro Arg Pro Val Arg Tyr Ala Ala Trp Ile Leu
100 105 110
Asn Glu Val Ala Met Met Ala Thr Asp Met Thr Gly Val Ile Gly Thr
115 120 125
Ala Ile Ala Leu Lys Met Leu Phe Gly Leu Pro Leu Leu Ala Gly Ile
130 135 140
Leu Leu Thr Ile Ala Asp Val Leu Val Val Leu Leu Phe Leu Arg Phe
145 150 155 160
Gly Ile Arg Arg Val Glu Val Ile Val Leu Val Ala Ile Leu Thr Val
165 170 175
Gly Ile Ile Phe Gly Ile Glu Val Gly Arg Ala His Val Gln Phe Gly
180 185 190
Asn Val Leu Leu Gly Leu Val Pro Thr Pro Leu Ile Val Lys Asn His
195 200 205
Thr Ala Leu Val Leu Ser Leu Gly Ile Leu Gly Ala Thr Ile Met Pro
210 215 220
His Asn Leu Tyr Leu His Ser Ser Leu Ala Gln Ser Arg Arg Tyr Asp
225 230 235 240
Tyr His Asn Pro Ala Gln Val Thr Glu Ala Leu Arg Phe Ala Asn Trp
245 250 255
Asp Ser Thr Val His Leu Ile Ala Ala Phe Leu Ile Asn Ala Leu Leu
260 265 270
Leu Val Leu Gly Gly Thr Leu Phe Phe Gly His Thr Asn Ala Leu Ala
275 280 285
Ser Leu Gln Ala Val Phe Asp Gly Leu Lys Ser Thr Thr Val Val Gly
290 295 300
Ala Leu Ala Ser Pro Val Met Ser Trp Leu Phe Ala Leu Ala Leu Leu
305 310 315 320
Ile Thr Gly Leu Ile Ser Ser Ile Thr Ser Thr Leu Ala Gly Gln Ile
325 330 335
Val Met Glu Gly Tyr Leu His Ile Arg Leu Pro Leu Trp Gln Arg Arg
340 345 350
Leu Leu Thr Arg Ala Val Thr Leu Ile Pro Ile Leu Ile Ile Gly Met
355 360 365
Leu Val Gly Phe Ser Asp Ala Ala Phe Glu Asn Leu Ile Ile Tyr Ala
370 375 380
Gln Val Ala Leu Ser Ile Ala Leu Pro Phe Thr Leu Leu Pro Leu Val
385 390 395 400
Ala Leu Thr Asn Asp Ala Ser Leu Met Lys Ala His Val Asn Arg Pro
405 410 415
Ala Val Thr Trp Val Gly Tyr Gly Leu Ala Gly Ile Ile Thr Val Leu
420 425 430
Asn Ile Tyr Leu Val Tyr Ser Leu Phe
435 440
<210> 17
<211> 434
<212> PRT
<213> 罗伊氏乳杆菌（Lactobacillus reuteri）
<400> 17
Met Glu Arg Lys Ser Leu Asp Glu Ile Asn Gly Ser Val Asp Val Pro
1 5 10 15
Asn Val Tyr Gln Ser Ala Phe Trp Gln Lys Phe Leu Ala Tyr Ser Gly
20 25 30
Pro Gly Ala Leu Val Ala Val Gly Tyr Met Asp Pro Gly Asn Trp Leu
35 40 45
Thr Ser Leu Ala Gly Gly Ser Gln Tyr Arg Tyr Gln Leu Leu Val Val
50 55 60
Leu Phe Thr Ala Ile Leu Ile Ala Met Tyr Met Gln Ser Leu Ala Ile
65 70 75 80
Lys Leu Gly Val Thr Thr Arg Thr Asp Leu Ala Gln Ala Ile Ala Arg
85 90 95
Arg Leu Pro Thr Pro Leu Arg Ile Ala Leu Trp Leu Phe Asn Glu Ile
100 105 110
Ala Met Met Ala Thr Asp Leu Thr Gly Val Val Gly Thr Ala Val Ala
115 120 125
Leu Asn Met Leu Phe Lys Leu Pro Leu Leu Ile Gly Val Leu Leu Thr
130 135 140
Ile Ala Asp Val Leu Val Val Leu Phe Phe Leu His Phe Gly Ile Arg
145 150 155 160
Arg Ile Glu Phe Ile Val Leu Thr Ala Ile Leu Val Val Gly Ala Ile
165 170 175
Phe Ala Ile Glu Val Cys Arg Ala His Pro Glu Phe Ser Ala Ile Met
180 185 190
Asp Gly Phe Val Pro Arg Ser Thr Ile Phe Thr Asn His Ser Glu Leu
195 200 205
Leu Ile Ser Leu Gly Ile Val Gly Ala Thr Ile Met Pro His Asn Ile
210 215 220
Tyr Leu His Ser Ser Leu Ala Gln Ser Arg Arg Tyr Asp Glu His Asp
225 230 235 240
Pro Lys Gln Val Lys Glu Thr Leu Arg Phe Ala Asn Trp Asp Ser Leu
245 250 255
Ile His Leu Phe Ala Ala Phe Ile Val Asn Ala Leu Leu Leu Ile Leu
260 265 270
Gly Gly Thr Leu Phe Phe His Ala Ala Ser Leu Gly Ser Leu Glu Asp
275 280 285
Val Phe Phe Gly Leu Lys Asn Pro Gln Ile Val Gly Ser Leu Ala Ser
290 295 300
Pro Leu Met Ser Trp Leu Phe Ala Phe Ala Leu Leu Val Thr Gly Leu
305 310 315 320
Ile Ser Ser Ile Thr Ser Thr Leu Ala Gly Gln Ile Val Met Glu Gly
325 330 335
Phe Ile Asn Ile Arg Leu Pro Leu Trp Lys Arg Arg Leu Leu Thr Arg
340 345 350
Ala Val Thr Leu Val Pro Ile Leu Ile Ile Gly Phe Met Ile Asn Phe
355 360 365
Lys Glu Glu Gln Phe Glu Gln Leu Ile Ile Tyr Ala Gln Ile Val Leu
370 375 380
Ser Ile Ala Leu Pro Phe Thr Leu Tyr Pro Leu Val Ala Leu Thr Gly
385 390 395 400
Asn Lys Lys Leu Met Gly Pro His Val Asn Ser Arg Trp Gln Thr Val
405 410 415
Leu Gly Tyr Ile Leu Ala Ser Leu Val Thr Gly Leu Asn Leu Leu Val
420 425 430
Leu Val
<210> 18
<211> 440
<212> PRT
<213> 壳状乳杆菌（Lactobacillus crustorum）
<400> 18
Met Thr Glu Lys Lys Ser Leu Asp Glu Ile Asn Gly Ser Val Ala Val
1 5 10 15
Pro Gln Tyr Asn Thr Ser Phe Phe Arg Lys Phe Leu Ala Tyr Ser Gly
20 25 30
Pro Gly Ala Leu Ile Ala Val Gly Tyr Met Asp Pro Gly Asn Trp Leu
35 40 45
Thr Ser Leu Val Gly Gly Ala His His Lys Tyr Gln Leu Leu Ser Val
50 55 60
Leu Leu Ile Ser Ile Leu Val Ala Thr Phe Met Gln Ser Leu Ser Ile
65 70 75 80
Arg Leu Gly Ile Ala Ser Arg Gln Asp Leu Ala Gln Ala Ile Ala Lys
85 90 95
Lys Ala Lys Lys Pro Val Arg Tyr Cys Leu Trp Ile Ile Asn Glu Leu
100 105 110
Ala Met Met Ala Thr Asp Leu Thr Gly Val Ile Gly Thr Ala Leu Ala
115 120 125
Leu Asn Met Leu Phe Lys Leu Pro Leu Val Phe Gly Val Leu Ile Thr
130 135 140
Ile Leu Asp Val Phe Leu Ile Leu Trp Phe Met Arg Phe Gly Ile Arg
145 150 155 160
Arg Ile Glu Ser Ile Val Val Ile Ser Ile Leu Thr Val Gly Leu Ile
165 170 175
Phe Ala Phe Glu Val Ser His Val Gln Pro Asn Leu Thr Ala Ile Phe
180 185 190
Lys Gly Phe Val Pro Ser Gln Thr Ile Ile Thr Asn Gln Asn Lys Leu
195 200 205
Ile Leu Ser Leu Gly Ile Ile Gly Ala Thr Ile Met Pro His Asn Ile
210 215 220
Tyr Leu His Ser Ala Leu Ala Gln Ser Arg Arg Tyr Asp Tyr His Asp
225 230 235 240
Ser Arg Gln Val Arg Glu Ala Leu Arg Phe Ala Asn Trp Asp Ser Ile
245 250 255
Val His Leu Ile Ala Ala Leu Ile Ile Asn Cys Leu Leu Leu Ile Leu
260 265 270
Gly Gly Thr Ile Phe Tyr Asp Lys Ala Asp Gln Leu Ala Ser Leu Met
275 280 285
Thr Val Phe Lys Gly Leu Met Asn Tyr Gln Val Val Gly Ser Leu Ala
290 295 300
Ser Ser Phe Met Ser Tyr Leu Phe Ala Phe Ala Leu Leu Val Thr Gly
305 310 315 320
Leu Ile Ser Ser Ile Thr Ser Thr Leu Ser Gly Gln Ile Val Met Glu
325 330 335
Gly Tyr Leu Asn Ile Arg Leu Pro Leu Trp Gln Arg Arg Leu Leu Thr
340 345 350
Arg Ile Ile Thr Leu Ile Pro Ile Leu Val Ile Gly Phe Leu Val His
355 360 365
Phe Asn Glu Val Ile Phe Glu Asp Leu Ile Val Tyr Ala Gln Ile Ala
370 375 380
Leu Ser Val Ala Leu Pro Phe Thr Leu Phe Pro Leu Val Tyr Leu Thr
385 390 395 400
Asn Asn Ala Lys Ile Met Gly Lys His Val Asn Lys Lys Trp Gln Thr
405 410 415
Ile Leu Gly Phe Val Leu Ala Leu Ile Ile Thr Ile Leu Asn Ile Val
420 425 430
Leu Ile Ala Thr Thr Leu Ser His
435 440
<210> 19
<211> 535
<212> PRT
<213> 干酪乳杆菌
<400> 19
Met Ser Asp Asp His Lys Lys Arg His Pro Ile Lys Leu Ile Gln Tyr
1 5 10 15
Ala Asn Gly Pro Ser Leu Glu Glu Ile Asn Gly Thr Val Glu Val Pro
20 25 30
His Gly Lys Gly Phe Trp Arg Thr Leu Phe Ala Tyr Ser Gly Pro Gly
35 40 45
Ala Leu Val Ala Val Gly Tyr Met Asp Pro Gly Asn Trp Ser Thr Ser
50 55 60
Ile Thr Gly Gly Gln Asn Phe Gln Tyr Leu Leu Ile Ser Val Ile Leu
65 70 75 80
Met Ser Ser Leu Ile Ala Met Leu Leu Gln Tyr Met Ala Ala Lys Leu
85 90 95
Gly Ile Val Ser Gln Met Asp Leu Ala Gln Ala Ile Arg Ala Arg Thr
100 105 110
Ser Lys Lys Leu Gly Ile Val Leu Trp Ile Leu Thr Glu Leu Ala Ile
115 120 125
Met Ala Thr Asp Ile Ala Glu Val Ile Gly Ala Ala Ile Ala Leu Tyr
130 135 140
Leu Leu Phe His Ile Pro Leu Val Ile Ala Val Leu Val Thr Val Leu
145 150 155 160
Asp Val Leu Val Leu Leu Leu Leu Thr Lys Ile Gly Phe Arg Lys Ile
165 170 175
Glu Ala Ile Val Val Ala Leu Ile Leu Val Ile Leu Leu Val Phe Val
180 185 190
Tyr Gln Val Ala Leu Ser Asp Pro Asn Met Gly Ala Leu Leu Lys Gly
195 200 205
Phe Ile Pro Thr Gly Glu Thr Phe Ala Ser Ser Pro Ser Ile Asn Gly
210 215 220
Met Ser Pro Ile Gln Gly Ala Leu Gly Ile Ile Gly Ala Thr Val Met
225 230 235 240
Pro His Asn Leu Tyr Leu His Ser Ala Ile Ser Gln Thr Arg Lys Ile
245 250 255
Asp His Lys Asn Pro Asp Asp Val Ala Gln Ala Val Lys Phe Ser Ala
260 265 270
Trp Asp Ser Asn Ile Gln Leu Ser Phe Ala Phe Val Val Asn Cys Leu
275 280 285
Leu Leu Val Met Gly Val Ala Val Phe Lys Ser Gly Ala Val Lys Asp
290 295 300
Pro Ser Phe Phe Gly Leu Phe Gln Ala Leu Ser Asp Ser Ser Thr Leu
305 310 315 320
Ser Asn Gly Val Leu Ile Ala Val Ala Lys Ser Gly Ile Leu Ser Ile
325 330 335
Leu Phe Ala Val Ala Leu Leu Ala Ser Gly Gln Asn Ser Thr Ile Thr
340 345 350
Gly Thr Leu Thr Gly Gln Val Ile Met Glu Gly Phe Val His Met Lys
355 360 365
Met Pro Leu Trp Ala Arg Arg Leu Val Thr Arg Ile Ile Ser Val Ile
370 375 380
Pro Val Ile Val Cys Val Met Leu Thr Ala Arg Asp Thr Pro Ile Gln
385 390 395 400
Gln His Glu Ala Leu Asn Thr Leu Met Asn Asn Ser Gln Val Phe Leu
405 410 415
Ala Phe Ala Leu Pro Phe Ser Met Leu Pro Leu Leu Met Phe Thr Asn
420 425 430
Ser Lys Val Glu Met Gly Asp Arg Phe Lys Asn Thr Gly Trp Val Lys
435 440 445
Val Leu Gly Trp Ile Ser Val Leu Gly Leu Thr Gly Leu Asn Leu Lys
450 455 460
Gly Leu Pro Asp Ser Ile Ala Gly Phe Phe Gly Asp His Pro Thr Ala
465 470 475 480
Thr Gln Thr Asn Met Ala Asn Ile Ile Ala Ile Val Leu Ile Val Ala
485 490 495
Ile Leu Ala Leu Leu Ala Trp Thr Ile Trp Asp Leu Tyr Lys Gly Asn
500 505 510
Gln Arg Tyr Glu Ala His Leu Ala Ala Val Ala Asp Glu Lys Glu Ala
515 520 525
Lys Ala Asp Val Asp Glu Gln
530 535
<210> 20
<211> 535
<212> PRT
<213> 鼠李糖乳杆菌
<400> 20
Met Ser Asp Asp His Lys Lys Lys His Ser Met Lys Leu Ile Gln Tyr
1 5 10 15
Ala Asn Gly Pro Ser Leu Glu Glu Ile Asn Gly Thr Val Glu Val Pro
20 25 30
His Gly Lys Gly Phe Trp Arg Thr Leu Phe Ala Tyr Ser Gly Pro Gly
35 40 45
Ala Leu Val Ala Val Gly Tyr Met Asp Pro Gly Asn Trp Ser Thr Ser
50 55 60
Ile Thr Gly Gly Gln Asn Phe Gln Tyr Leu Leu Ile Ser Val Ile Leu
65 70 75 80
Met Ser Ser Leu Ile Ala Met Leu Leu Gln Tyr Met Ala Ala Lys Leu
85 90 95
Gly Ile Val Ser Gln Met Asp Leu Ala Gln Ala Ile Arg Ala Arg Thr
100 105 110
Ser Lys Lys Leu Gly Ile Val Leu Trp Ile Leu Thr Glu Leu Ala Ile
115 120 125
Met Ala Thr Asp Ile Ala Glu Val Ile Gly Ala Ala Ile Ala Leu Tyr
130 135 140
Leu Leu Phe His Ile Pro Leu Val Ile Ala Val Leu Val Thr Val Leu
145 150 155 160
Asp Val Leu Val Leu Leu Leu Leu Thr Lys Ile Gly Phe Arg Lys Ile
165 170 175
Glu Ala Ile Val Val Ala Leu Ile Leu Val Ile Leu Leu Val Phe Val
180 185 190
Tyr Gln Val Ala Leu Ser Asp Pro Asn Met Gly Ala Leu Leu Lys Gly
195 200 205
Phe Ile Pro Thr Gly Glu Thr Phe Ala Ser Ser Pro Ser Val Asn Gly
210 215 220
Met Ser Pro Ile Gln Gly Ala Leu Gly Ile Ile Gly Ala Thr Val Met
225 230 235 240
Pro His Asn Leu Tyr Leu His Ser Ala Ile Ser Gln Thr Arg Lys Ile
245 250 255
Asp His Lys Asp Pro Glu Asp Val Ala Gln Ala Val Lys Phe Ser Ala
260 265 270
Trp Asp Ser Asn Ile Gln Leu Thr Phe Ala Phe Val Val Asn Cys Leu
275 280 285
Leu Leu Val Met Gly Val Ala Val Phe Lys Ser Gly Ala Val Lys Asp
290 295 300
Pro Ser Phe Phe Gly Leu Phe Gln Ala Leu Ser Asp Ser Ser Thr Leu
305 310 315 320
Ser Asn Gly Val Leu Ile Ala Val Ala Lys Ser Gly Ile Leu Ser Ile
325 330 335
Leu Phe Ala Val Ala Leu Leu Ala Ser Gly Gln Asn Ser Thr Ile Thr
340 345 350
Gly Thr Leu Thr Gly Gln Val Ile Met Glu Gly Phe Ile His Met Lys
355 360 365
Met Pro Leu Trp Ala Arg Arg Leu Val Thr Arg Val Ile Ser Val Ile
370 375 380
Pro Val Ile Val Cys Val Met Leu Thr Ala Arg Glu Thr Pro Ile Gln
385 390 395 400
Gln His Glu Ala Leu Asn Thr Leu Met Asn Asn Ser Gln Val Phe Leu
405 410 415
Ala Phe Ala Leu Pro Phe Ser Met Leu Pro Leu Leu Met Phe Thr Asn
420 425 430
Ser Lys Val Glu Met Gly Asp Arg Phe Lys Asn Thr Gly Trp Val Lys
435 440 445
Val Leu Gly Trp Val Ser Val Ile Gly Leu Thr Tyr Leu Asn Leu Lys
450 455 460
Gly Leu Pro Asp Ser Ile Ala Gly Phe Phe Gly Asp Asn Pro Thr Ala
465 470 475 480
Ala Gln Thr Asn Ile Ala Asn Met Ile Ala Tyr Val Leu Ile Ala Ala
485 490 495
Val Leu Ala Leu Leu Ala Trp Thr Ile Trp Asp Leu Tyr Lys Gly Asn
500 505 510
Lys Arg Tyr Glu Ala His Leu Glu Ala Val Ala Asp Glu Glu Glu Ala
515 520 525
Lys Ala Asn Asp Asp Val Gln
530 535
<210> 21
<211> 530
<212> PRT
<213> 植物乳杆菌
<400> 21
Met Ser Glu Lys Thr Asn Thr Pro Asn Arg Lys His Lys Leu Ile Glu
1 5 10 15
Tyr Ala Asn Gly Pro Ser Leu Glu Glu Ile Asn Gly Thr Ile Glu Val
20 25 30
Pro Lys Asn Leu Asn Phe Trp Lys Thr Leu Phe Ala Tyr Ser Gly Pro
35 40 45
Gly Ala Leu Val Ala Val Gly Tyr Met Asp Pro Gly Asn Trp Ser Thr
50 55 60
Ser Ile Thr Gly Gly Gln Asn Tyr Gln Tyr Met Leu Met Ser Val Ile
65 70 75 80
Leu Ile Ser Ser Leu Ile Ala Met Leu Leu Gln Tyr Met Ala Ala Lys
85 90 95
Leu Gly Ile Val Ser Gln Met Asp Leu Ala Gln Ala Ile Arg Ala Arg
100 105 110
Thr Ser Lys Ser Leu Gly Ile Val Leu Trp Ile Leu Thr Glu Leu Ala
115 120 125
Ile Met Ala Thr Asp Ile Ala Glu Val Ile Gly Ala Ala Ile Ala Leu
130 135 140
Tyr Leu Leu Phe Asn Ile Pro Leu Val Ile Ala Val Phe Ile Thr Val
145 150 155 160
Leu Asp Val Leu Val Leu Leu Leu Leu Thr Lys Ile Gly Phe Arg Lys
165 170 175
Ile Glu Ala Ile Val Val Cys Leu Ile Leu Val Ile Leu Phe Val Phe
180 185 190
Val Tyr Gln Val Ala Leu Ser Asn Pro Asp Trp Gly Gly Val Ile Lys
195 200 205
Gly Leu Val Pro Thr Ala Asp Thr Phe Ser Thr Ser Arg Ser Val Asn
210 215 220
Gly Met Thr Pro Leu Ser Gly Ala Leu Gly Ile Ile Gly Ala Thr Val
225 230 235 240
Met Pro His Asn Leu Tyr Leu His Ser Ala Ile Ser Gln Thr Arg Lys
245 250 255
Ile Asp His Asn Asp Glu Glu Asp Val Ala Arg Thr Val Lys Phe Ala
260 265 270
Ala Trp Asp Ser Asn Ile Gln Leu Ser Phe Ala Phe Val Val Asn Ser
275 280 285
Leu Leu Leu Ile Met Gly Val Ala Val Phe Lys Ser Gly Ala Val Lys
290 295 300
Asp Pro Ser Phe Phe Gly Leu Tyr Glu Ala Leu Ser Asn Thr Ser Met
305 310 315 320
Leu Ser Asn Gly Ile Leu Ile Ser Val Ala Lys Ser Gly Ala Leu Ser
325 330 335
Ala Leu Phe Ala Ile Ala Leu Leu Ala Ser Gly Gln Asn Ser Thr Ile
340 345 350
Thr Gly Thr Leu Thr Gly Gln Val Ile Met Glu Gly Phe Val His Met
355 360 365
Arg Met Pro Leu Trp Leu Arg Arg Leu Val Thr Arg Leu Ile Ser Val
370 375 380
Ile Pro Val Leu Ile Cys Val Leu Leu Thr Ser Gly Lys Ser Ala Ile
385 390 395 400
Asp Glu His Thr Ala Leu Asn Asn Leu Met Asn Asn Ser Gln Val Phe
405 410 415
Leu Ala Phe Ala Leu Pro Phe Ser Met Leu Pro Leu Leu Met Met Thr
420 425 430
Asp Ser Ala Ala Glu Met Gly Lys Arg Phe Lys Asn Ser Leu Trp Ile
435 440 445
Lys Gly Leu Gly Trp Leu Ser Val Ile Gly Leu Thr Phe Leu Asn Leu
450 455 460
Leu Gly Leu Pro Asp Ser Ile Leu Gly Phe Phe Gly Asp Asn Pro Ser
465 470 475 480
Ala Gly Glu Gln Thr Phe Ser Lys Ile Leu Ala Tyr Leu Leu Ile Ala
485 490 495
Ala Ile Leu Ala Leu Leu Val Trp Thr Val Phe Asp Leu Gln Arg Gly
500 505 510
Asn Lys Arg Tyr Val Glu Gln Gln Leu Ala Ala Ala Ala Lys Glu Ala
515 520 525
Asn Lys
530
<210> 22
<211> 530
<212> PRT
<213> 乳酸片球菌
<400> 22
Met Ser Asn Glu Ile Lys Asn Pro Lys Lys Arg Arg Lys Leu Ile Ser
1 5 10 15
Tyr Ala Asn Gly Arg Ser Leu Glu Glu Ile Asn Gly Thr Val Lys Val
20 25 30
Pro Lys Asn Ile Gly Phe Trp Lys Thr Leu Phe Met Tyr Ser Gly Pro
35 40 45
Gly Ala Leu Val Ala Val Gly Tyr Met Asp Pro Gly Asn Trp Ser Thr
50 55 60
Ser Ile Thr Gly Gly Gln Asn Phe Gln Tyr Met Leu Met Ser Ile Ile
65 70 75 80
Leu Ile Ser Ser Leu Ile Ala Met Leu Leu Gln Tyr Met Ala Ala Lys
85 90 95
Leu Gly Ile Val Ser Gln Met Asp Leu Ala Gln Ala Ile Arg Ala Arg
100 105 110
Thr Ser Arg Ala Leu Gly Ile Val Leu Trp Ile Leu Thr Glu Leu Ala
115 120 125
Ile Met Ala Thr Asp Ile Ala Glu Val Ile Gly Ala Ala Ile Ala Leu
130 135 140
Tyr Leu Leu Phe His Ile Pro Leu Val Val Ala Val Phe Ile Thr Val
145 150 155 160
Phe Asp Val Leu Leu Leu Leu Leu Leu Thr Lys Ile Gly Phe Arg Lys
165 170 175
Ile Glu Ala Ile Val Val Cys Leu Ile Met Val Ile Leu Val Val Phe
180 185 190
Val Tyr Gln Val Ala Leu Ser His Pro Ser Trp Gly Ala Val Phe Gly
195 200 205
Gly Leu Ile Pro Thr Thr Lys Ala Phe Ala Thr Thr Pro Thr Val Gly
210 215 220
Gly Met Thr Pro Leu Ser Gly Ser Leu Gly Ile Ile Gly Ala Thr Val
225 230 235 240
Met Pro His Asn Leu Tyr Leu His Ser Ala Val Ser Gln Thr Arg Lys
245 250 255
Ile Asn His Asp Asp Glu Glu Asp Val Ala Arg Thr Val Arg Phe Ser
260 265 270
Thr Trp Asp Ser Asn Ile Gln Leu Ser Phe Ala Phe Val Val Asn Ala
275 280 285
Leu Leu Leu Val Met Gly Val Ala Val Phe Lys Thr Gly Ala Val Gln
290 295 300
Asp Pro Ser Phe Phe Gly Leu Phe His Ala Leu Asn Asp Thr Ser Thr
305 310 315 320
Leu Ser Asn Gly Ile Leu Ile Gly Val Ala Lys Thr Gly Ile Leu Ser
325 330 335
Thr Leu Phe Ala Val Ala Leu Leu Ala Ser Gly Gln Asn Ser Thr Ile
340 345 350
Thr Gly Thr Leu Thr Gly Gln Val Ile Met Glu Gly Phe Val His Met
355 360 365
Arg Met Pro Leu Trp Ala Arg Arg Leu Ile Thr Arg Leu Ile Ser Val
370 375 380
Val Pro Val Leu Ile Cys Val Met Leu Thr Ser Gly Lys Gly Thr Ile
385 390 395 400
Gln Glu His Glu Ala Leu Asn Asn Leu Met Asn Asn Ser Gln Val Phe
405 410 415
Leu Ala Phe Ala Leu Pro Phe Ser Met Val Pro Leu Leu Met Met Thr
420 425 430
Asp Ser Arg Val Glu Met Gly Asp Arg Phe Lys Asn Ser Trp Ile Val
435 440 445
Arg Ile Leu Gly Trp Ile Ser Val Ile Phe Leu Thr Tyr Leu Asn Leu
450 455 460
Thr Gly Leu Pro Asp Ser Ile Ala Ala Phe Phe Gly Glu Asn Ala Ser
465 470 475 480
Ala Ala Glu Ile Ser Met Ala His Asp Ile Ala Tyr Ala Leu Ile Val
485 490 495
Ala Val Leu Ala Leu Leu Ala Trp Thr Val Ile Glu Leu Tyr Lys Gly
500 505 510
Asn Lys Arg Tyr Glu Ile Glu Leu Ala Glu Lys Ala Asn Ala Lys Glu
515 520 525
Ala Ala
530
<210> 23
<211> 521
<212> PRT
<213> 唾液乳杆菌（Lactobacillus salivarius）
<400> 23
Met Val Asn Asn Glu Asn Asn His Lys Lys His Lys Met Ile Gln Tyr
1 5 10 15
Ala Asn Gly Lys Ser Leu Glu Glu Ala Asn Gly Thr Val Glu Ile Pro
20 25 30
Lys Gly Lys Gly Phe Trp Lys Thr Leu Phe Ala Tyr Ser Gly Pro Gly
35 40 45
Ala Leu Val Ala Val Gly Tyr Met Asp Pro Gly Asn Trp Ser Thr Ser
50 55 60
Ile Thr Gly Gly Gln Asn Phe Gln Tyr Leu Leu Met Ser Val Ile Leu
65 70 75 80
Leu Ser Ser Leu Ile Ala Met Leu Leu Gln Tyr Met Ala Ala Lys Leu
85 90 95
Gly Ile Val Ser Gln Met Asp Leu Ala Gln Ala Ile Arg Ala Arg Thr
100 105 110
Ser Lys Ala Leu Gly Ile Val Leu Trp Ile Leu Thr Glu Leu Ala Ile
115 120 125
Met Ala Thr Asp Ile Ala Glu Val Ile Gly Ala Ala Ile Ala Leu Tyr
130 135 140
Leu Leu Phe Asp Ile Pro Leu Ile Ile Ala Val Phe Ile Thr Val Phe
145 150 155 160
Asp Val Leu Leu Leu Leu Leu Leu Thr Lys Val Gly Phe Arg Lys Ile
165 170 175
Glu Ala Ile Val Val Cys Leu Ile Phe Val Ile Leu Phe Val Phe Val
180 185 190
Tyr Gln Val Ala Leu Ser Asn Pro Asp Trp Gly Gly Val Phe Lys Gly
195 200 205
Leu Ile Pro Thr Ser Glu Thr Phe Ala Lys His Pro Val Val His Asp
210 215 220
Met Ser Pro Leu Asn Gly Ala Leu Gly Ile Ile Gly Ala Thr Val Met
225 230 235 240
Pro His Asn Leu Tyr Leu His Ser Ala Ile Ser Gln Thr Arg Lys Phe
245 250 255
Asp Arg Asn Asn Glu Asp Asp Ile Ala Asn Ala Val Arg Phe Thr Ala
260 265 270
Trp Asp Ser Asn Ile Gln Leu Gly Leu Ala Phe Val Val Asn Ser Leu
275 280 285
Leu Leu Ile Met Gly Val Ala Val Phe Lys Ser Gly Ala Val Glu Asp
290 295 300
Pro Ser Phe Phe Gly Leu Tyr Gln Ala Leu Ser Asp Thr Ser Val Met
305 310 315 320
Ser Asn Gly Leu Leu Ala Ala Ala Ala Arg Thr Gly Ile Leu Ser Thr
325 330 335
Leu Phe Ala Val Ala Leu Leu Ala Ser Gly Gln Asn Ser Thr Ile Thr
340 345 350
Gly Thr Leu Thr Gly Gln Val Ile Met Glu Gly Phe Ile His Leu Arg
355 360 365
Met Pro Leu Trp Ala Arg Arg Leu Ile Thr Arg Leu Leu Ser Val Ile
370 375 380
Pro Val Leu Ile Cys Val Ala Leu Thr Ser Gly Lys Ser Thr Ile Glu
385 390 395 400
Glu His Glu Ala Leu Asn Asn Leu Met Asn Asn Ser Gln Val Phe Leu
405 410 415
Ala Phe Ala Leu Pro Phe Ser Met Leu Pro Leu Val Ile Met Thr Gly
420 425 430
Ser Lys Val Glu Met Gly Glu Arg Phe Lys Asn Arg Leu Trp Ile Asn
435 440 445
Ile Leu Gly Trp Ile Ser Val Ile Ser Leu Thr Tyr Leu Asn Met Ile
450 455 460
Gly Leu Pro Gln Asn Leu Glu Pro Phe Phe Pro Ala Asp Lys Val Gly
465 470 475 480
Leu Ala His Thr Val Ala Tyr Ile Leu Ile Val Leu Ile Ile Ala Leu
485 490 495
Leu Ile Trp Thr Leu Val Glu Leu His Leu Gly Asn Lys Arg Phe Ala
500 505 510
Ala Glu Gln Ala Lys Lys His Asn Lys
515 520
<210> 24
<211> 520
<212> PRT
<213> 发酵乳杆菌（Lactobacillus fermentum）
<400> 24
Met Asp Asn Thr Lys Asn Gln His Arg Lys Leu Arg Leu Ile Glu His
1 5 10 15
Ala Asn Gly Lys Ser Leu Glu Glu Ile Asn Gly Thr Val Glu Val Pro
20 25 30
His Gly Lys Gly Phe Phe Arg Thr Leu Phe Ala Tyr Ser Gly Pro Gly
35 40 45
Ala Leu Val Ala Val Gly Tyr Met Asp Pro Gly Asn Trp Ser Thr Ser
50 55 60
Ile Thr Gly Gly Gln Ser Phe Gln Tyr Thr Leu Met Thr Thr Ile Leu
65 70 75 80
Ile Ser Ser Leu Ile Ala Met Leu Leu Gln Tyr Met Ala Ala Lys Leu
85 90 95
Gly Ile Val Ser Gln Met Asp Leu Ala Gln Ala Ile Arg Ala Arg Thr
100 105 110
Gly Lys Ala Leu Gly Val Ile Leu Trp Leu Met Thr Glu Leu Ala Ile
115 120 125
Met Ala Thr Asp Ile Ala Glu Val Ile Gly Ala Ala Ile Ala Leu Asn
130 135 140
Leu Leu Phe His Ile Pro Leu Val Leu Ala Val Phe Ile Thr Val Leu
145 150 155 160
Asp Val Leu Val Leu Leu Leu Leu Thr Lys Ile Gly Phe Arg Lys Ile
165 170 175
Glu Ala Ile Val Ala Cys Leu Ile Leu Val Ile Leu Ala Val Phe Ala
180 185 190
Tyr Gln Val Ala Leu Ser His Pro Asp Trp Ala Gly Val Phe Lys Gly
195 200 205
Leu Leu Pro Thr Lys Glu Ala Ile Ala Lys Glu Pro Val Val Gly Gly
210 215 220
Ile Ser Pro Leu Thr Gly Ser Leu Gly Ile Ile Gly Ala Thr Val Met
225 230 235 240
Pro His Asn Leu Tyr Leu His Ser Ala Ile Ser Gln Thr Arg Lys Ile
245 250 255
Asp His Thr Asn Ala Glu Asp Ile Lys Gln Thr Val Arg Phe Thr Ala
260 265 270
Trp Asp Ser Asn Ile Gln Leu Thr Leu Ala Phe Phe Val Asn Ala Leu
275 280 285
Leu Leu Ile Met Gly Val Ala Val Phe Lys Asn Gly Ala Val Gln Asp
290 295 300
Ser Ser Phe Phe Gly Leu Tyr Asp Ala Leu Asn Asn Thr Asp Met Leu
305 310 315 320
Ser Asn Gly Leu Leu Ile Ala Val Ala Lys Ser Gly Val Leu Ser Thr
325 330 335
Leu Phe Ala Ile Ala Leu Leu Ala Ser Gly Gln Asn Ser Thr Ile Thr
340 345 350
Gly Thr Leu Thr Gly Gln Val Ile Met Glu Gly Phe Val His Met Lys
355 360 365
Met Pro Leu Trp Ala Arg Arg Leu Ile Thr Arg Leu Leu Ser Val Val
370 375 380
Pro Val Leu Val Cys Val Ala Met Thr Ala His Glu Ser Thr Ile Asp
385 390 395 400
Gln His Ala Ser Leu Asn Ile Leu Met Glu Asn Ser Gln Val Phe Leu
405 410 415
Ala Phe Ala Leu Pro Phe Ser Met Leu Pro Leu Leu Ile Met Thr Asn
420 425 430
Ser Asp Thr Glu Met Gly Gln Phe Lys Asn Ser Leu Trp Val Arg Val
435 440 445
Leu Gly Trp Ile Ser Val Ile Gly Leu Thr Phe Leu Asn Leu Tyr Asn
450 455 460
Leu Pro Gln Thr Tyr Glu Gly Phe Gly Ile Trp Ser Lys Gly Leu Ser
465 470 475 480
Asp Val Leu Ala Trp Ile Ser Ile Val Val Ile Val Val Leu Leu Ala
485 490 495
Trp Thr Cys Phe Glu Leu Ile Arg Gly Asp Arg Arg Leu Ala Ala Glu
500 505 510
Arg Glu Lys His Thr Trp Glu Lys
515 520
<210> 25
<211> 533
<212> PRT
<213> 解淀粉乳杆菌（Lactobacillus amylolyticus）
<400> 25
Met Cys Ser Arg Lys Val Leu Leu Thr Lys Gln Lys Gly Lys His Tyr
1 5 10 15
Leu Ile Arg Tyr Ala Asn Gly Lys Ser Leu Ser Glu Ile Asn Gly Thr
20 25 30
Ile Glu Ile Pro Lys Lys Arg Thr Phe Trp Arg Met Leu Trp Ala Tyr
35 40 45
Thr Gly Pro Gly Ala Leu Val Ala Val Gly Tyr Met Asp Pro Gly Asn
50 55 60
Trp Ala Thr Ser Ile Thr Gly Gly Gln Ser Phe Gln Tyr Ile Leu Met
65 70 75 80
Ser Thr Ile Leu Ile Ser Ser Leu Met Ala Met Leu Leu Gln Tyr Met
85 90 95
Ala Ala Lys Leu Gly Ile Val Thr Gln Met Asp Leu Ala Gln Ala Ile
100 105 110
Arg Leu Arg Thr Gly Lys Ala Leu Gly Ile Val Leu Trp Leu Met Thr
115 120 125
Glu Leu Ala Ile Met Ala Thr Asp Ile Ala Glu Val Ile Gly Ala Ala
130 135 140
Ile Ala Leu Asn Leu Leu Phe Asp Ile Pro Leu Val Pro Ala Val Phe
145 150 155 160
Ile Thr Val Leu Asp Val Leu Leu Leu Leu Leu Leu Ala Arg Ile Gly
165 170 175
Phe Arg Lys Ile Glu Ala Val Val Ser Cys Leu Ile Leu Val Ile Leu
180 185 190
Leu Val Phe Val Tyr Glu Val Leu Leu Ser Asn Pro Asp Trp Ser Lys
195 200 205
Ala Phe Val Gly Leu Val Pro Ser Ala Lys Ile Ile Gln Thr His Pro
210 215 220
Val Val Gly Gly Ile Ser Pro Leu Thr Gly Thr Leu Gly Ile Ile Gly
225 230 235 240
Ala Thr Val Met Pro His Asn Leu Tyr Leu His Ser Ala Ile Ser Gln
245 250 255
Thr Arg Lys Ile Asn His His Asn Leu Gln Leu Ile Arg Asp Ala Val
260 265 270
Lys Tyr Thr Ala Leu Asp Ser Asn Ile Gln Leu Ser Leu Ala Phe Leu
275 280 285
Val Asn Ala Leu Leu Leu Ile Met Gly Ala Ala Val Phe Lys Ser Gly
290 295 300
Ala Val Arg Asp Ser Ser Phe Phe Gly Leu Tyr Gln Ala Leu Asp Asn
305 310 315 320
Ala Lys Met Leu Ser Asp Pro Leu Leu Val His Val Ala Arg Thr Gly
325 330 335
Ile Leu Ser Thr Leu Phe Ala Val Ala Leu Leu Ala Ser Gly Gln Asn
340 345 350
Ser Thr Ile Thr Gly Thr Leu Thr Gly Gln Val Ile Met Glu Gly Tyr
355 360 365
Ile His Leu Lys Met Pro Leu Trp Ala Arg Arg Leu Val Thr Arg Leu
370 375 380
Leu Ser Val Ile Pro Val Leu Leu Cys Val Ser Phe Thr Met Asn Asp
385 390 395 400
Ser Val Met Gln Gln His Phe Ala Leu Asn Met Leu Met Glu Asn Ser
405 410 415
Gln Val Phe Leu Ala Phe Ala Leu Pro Phe Ser Val Leu Pro Leu Leu
420 425 430
Ile Met Thr Asn Asn Lys Ala Glu Met Gly Glu Phe Lys Asn Lys Pro
435 440 445
Leu Trp His Tyr Leu Gly Trp Ala Cys Ala Leu Val Leu Thr Phe Leu
450 455 460
Asn Leu Tyr Asn Leu Pro Ser Gln Phe Val Asn Phe Lys Phe Ala Ser
465 470 475 480
Lys Glu Val Ser Thr Ile Ile Ala Tyr Phe Val Ile Val Val Ile Ala
485 490 495
Ala Leu Leu Leu Trp Thr Cys Ile Glu Ile Tyr Ile Gly Asp Arg Lys
500 505 510
Val Lys Ile His His Ser Gly Phe Asp Ala Lys Glu Lys Glu Leu Lys
515 520 525
Glu Glu Gly Gln Lys
530

Claims (15)

1.一种包含含锰转运蛋白的乳酸菌的直投式起子培养物组合物，其用于发酵食物产品和用于抑制或延迟所述食物产品中的真菌生长，
其特征在于
所述组合物包含至多600ppm的锰并且
所述乳酸菌的浓度为至少1E+10菌落形成单位/g，
其中，任选地，所述乳酸菌不含超氧化物歧化酶，优选不含锰超氧化物歧化酶。
2.根据前一权利要求所述的组合物，其包含至多400ppm的锰，优选至多300ppm的锰，更优选至多250ppm的锰，甚至更优选至多200ppm的锰。
3.根据前述权利要求中任一项所述的组合物，其包含30-600ppm、优选35-600ppm或40-400ppm或40-300ppm或40-250ppm的锰，更优选包含40-200ppm或45-200ppm的锰。
4.根据前述权利要求中任一项所述的组合物，其中所述乳酸菌的浓度为2.0E+10-6.5E+11菌落形成单位/g，优选6.0E+10-6.4E+11菌落形成单位/g，更优选1.3E+11-5.6E+11菌落形成单位/g。
5.根据前述权利要求中任一项所述的组合物，其中所述组合物是冷冻直投式(F-DVS)或冷冻干燥直投式(FD-DVS)。
6.根据前述权利要求中任一项所述的组合物，其中所述组合物是冷冻干燥直投式(FD-DVS)组合物，其包含以每重量FD-DVS形式的添加剂干重测量的2％到70％的添加剂、优选3％到50％的添加剂、更优选4％到40％的添加剂、甚至更优选10％到30％的添加剂或20％到30％的添加剂，优选其中所述添加剂不含锰或基本上不含锰，或其中所述组合物是冷冻直投式(F-DVS)，其包含以每重量所述F-DVS形式的添加剂重量测量的2％到70％的添加剂、优选3％到50％的添加剂、更优选4％到40％的添加剂、甚至更优选10％到40％的添加剂或20％至35％的添加剂，优选其中所述添加剂不含锰或基本上不含锰。
7.根据前一权利要求所述的组合物，其选自由以下组成的组：酪蛋白酸钠、肌醇、谷氨酸单钠、抗坏血酸钠、蔗糖、麦芽糖糊精、肌苷一磷酸(IMP)、肌苷、聚山梨糖醇酯80、谷氨酸、赖氨酸、谷氨酸钠、麦芽提取物、乳清粉、酵母提取物、麸质、胶原蛋白、明胶、弹性蛋白、角蛋白、白蛋白和其混合物。
8.根据前一权利要求所述的组合物，其包含酪蛋白酸钠、肌醇、谷氨酸单钠和抗坏血酸钠。
9.根据前述权利要求中任一项所述的组合物，其中所述乳酸菌包含锰转运蛋白，所述锰转运蛋白与SEQ ID NO:1-3中任一个的序列具有至少55％，诸如至少60％、65％、70％、75％、80％、85％、90％、95％、96％、97％、98％、99％或100％的序列同一性。
10.根据前述权利要求中任一项所述的组合物，其中所述乳酸菌不含超氧化物歧化酶，优选不含锰超氧化物歧化酶。
11.根据前述权利要求中任一项所述的组合物，其中所述乳酸菌选自由以下组成的组：植物乳杆菌(Lactobacillus plantarum)、发酵乳杆菌(Lactobacillus fermentum)、罗伊氏乳杆菌(Lactobacillus reuteri)、沙克乳杆菌(Lactobacillus sakei)、短乳杆菌(Lactobacillus brevis)、干酪乳杆菌(Lactobacillus casei)、副干酪乳杆菌(Lactobacillus paracasei)、唾液乳杆菌(Lactobacillus salivarius)、消化乳杆菌(Lactobacillus alimentarius)、乳酸片球菌(Pediococcus acidilactici)、鼠李糖乳杆菌(Lactobacillus rhamnosus)和开菲尔乳杆菌(Lactobacillus kefiri)。
12.根据前述权利要求中任一项所述的组合物，其中所述真菌是酵母和/或霉菌，优选其中所述真菌是选自由以下组成的组的酵母：有孢圆酵母属的种(Torulaspora spp.)、隐球菌属的种(Cryptococcus spp.)、酵母属的种(Saccharomyces spp.)、耶氏酵母属的种(Yarrowia spp.)、德巴利酵母属的种(Debaryomyces spp.)、假丝酵母属的种(Candidaspp.)和红酵母属(Rhodoturola)，优选其中德巴利酵母属的种是汉逊德巴利酵母(Debaromyces hansenii)和/或其中所述真菌是选自由以下组成的组的霉菌：曲霉属的种(Aspergillus spp.)、枝孢菌属的种(Cladosporium spp.)、亚隔孢壳属的种(Didymellaspp.)或青霉属的种(Penicillium spp.)，优选其中青霉属的种是皮壳青霉(Penicilliumcrustosum)、潘氏青霉(Penicillium paneum)、卡尼青霉(Penicillium carneum)或娄地青霉(Penicillium roqueforti)。
13.一种食物产品，其包含根据前述权利要求中任一项所述的组合物。
14.根据前一权利要求所述的食物产品，其中所述食物产品是发酵食物产品，优选嗜热发酵食物产品或嗜中温发酵食物产品，更优选酸奶或奶酪。
15.根据前述权利要求1-12中任一项所述的组合物用于抑制食物产品中真菌生长的用途，优选其中所述食物产品是发酵食物产品，更优选嗜热发酵食物产品或嗜中温发酵食物产品，更优选酸奶或奶酪。
CN202080073495.4A 2019-10-23 2020-10-21 用于控制真菌腐败的细菌组合物和其用途 Pending CN114585258A (zh)

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