CN1219106A - 一种获得蛋白质水解物的方法 - Google Patents
一种获得蛋白质水解物的方法 Download PDFInfo
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- CN1219106A CN1219106A CN97194838A CN97194838A CN1219106A CN 1219106 A CN1219106 A CN 1219106A CN 97194838 A CN97194838 A CN 97194838A CN 97194838 A CN97194838 A CN 97194838A CN 1219106 A CN1219106 A CN 1219106A
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Abstract
本发明涉及一种获得用作调味剂的蛋白质水解物的方法。更特别的,本发明提供了从蛋白质底物获得富含游离谷氨酸和/或肽结合的谷氨酸残基的水解物的一种方法,该方法包括将该底物进行脱酰胺处理且将该底物经过一种特异性作用蛋白水解酶的作用的步骤。
Description
本发明涉及一种获得用作调味剂的蛋白质水解物的方法。特别是,本发明提供一种从蛋白质底物获得富含游离谷氨酸和/或肽结合的谷氨酸残基的水解物的方法,该方法包括将该底物经过一种脱酰胺过程,且将该底物经过一种特异性作用蛋白质水解酶的作用的步骤。
不同的食物产品,如汤、调味汁和调味品,包含通过蛋白质物质水解获得的调味剂。传统地,这种水解是利用强盐酸后再用氢氧化钠中和的方法进行。然而,这种化学水解法导致在水解过程中获得的氨基酸的严重降解,且在此化学反应过程中形成一些危险的副产物。因此,对通过化学水解所获得调味剂的使用的日益担心导致了酶法水解工艺的发展。
酶法水解工艺以获得高水解度(DH)为目的,且通常使用非特异性作用蛋白水解酶复合物(即:非特异性作用内肽酶和外肽酶)来实现。在这方面,如WO 94/25580描述了使用一种获自米曲霉(Aspergillusoryzae)的非特异性作用酶制剂水解蛋白质的一种方法。特异性作用蛋白水解酶尚未被用于此目的,因为这种酶仅导致一种不充分的水解。
目前仅报道了几种特异性作用蛋白水解酶。例如,已报道了优选地在谷氨酰肽键(谷氨酸特异性蛋白酶)处切割肽的蛋白水解酶获自金黄色葡萄球菌(Staphylococcus aureus)[Drapeau G.R;Boily Y.和Houmard J;生物化学杂志(J.Biol.Chem.)1972,247(20)6720-6726],获自放线菌属(Actinomyces sp.)[Mosolova O.V.,Rudenskaya G.N.,Stepanov V.M.,Khodova O.M.和Tsaplina I.A.;Biokhimiya 1987,52(3)414-422],获自灰色链霉菌(Streptomyces griseus)[Yoshida N.,Tsuruyama S.,Nagata K,Hirayama K,Noda K.和Makisumi S.;生物化学杂志(J.Biochem.)1988,104,451-456],获自热普通链霉菌(Streptomyces thermovulgaris)[Khaldarova N.Y.,Rudenskaya G.N.,Revina L.P.,Stephanov V.M.和Egorov N.S.;Biochimiia Moscow Eng.1989,54(1)32-38],获自枯草芽孢杆菌(Bacillus subtilis)[Niidome T.,Yoshida N.,Ogata F.,lto A.和Noda K.;生物化学杂志(J.Biochem.),1990,108,965-970]以及获自地衣芽孢杆菌(Bacillus licheniforms)[WO91/13554 A1]。
谷氨酸特异性蛋白酶最初应用于蛋白质结构研究。然而,已报道了谷氨酸特异性金黄色葡萄球菌蛋白酶用于修饰酪蛋白的溶解性和结构性质的用途[Chobert J-M.,Sitohy M.Z.和Whitaker J.R.;农业食品化学杂志(J.Agric.Food Chem.)1988,36,220-224],以及WO 91/13554A1描述了地衣芽孢杆菌用于获得蛋白质有限的特异性水解的用途。
虽然谷氨酰胺(Gln)几乎没有味道,谷氨酸(Glu)无论是游离的还是肽结合的,均对蛋白质水解产物的香味和口味起重要作用。谷氨酸可以通过将游离谷氨酰胺转化为谷氨酸来形成。这种转化(即脱酰胺)在使用强盐酸的传统化学水解过程中自然发生。
此外,谷氨酸可以通过谷氨酰胺酶(L-谷氨酰胺酶,EC3.5.1.22或D-谷氨酰胺酶,EC3.5.1.35)的作用从游离的谷氨酰胺形成。然而,由于其自发地转变成焦谷氨酰胺,大量谷氨酰胺会损失掉。
与谷氨酰胺酶相反,肽聚谷氨酰胺酶(peptidoglutaminase)(PG酶)作用于肽结合的谷氨酰胺。已知两种类型的肽聚谷氨酰胺酶。肽酰谷氨酰胺酶(EC3.5.1.43;肽聚谷氨酰胺酶Ⅰ)特异于在α-氨基上有取代的谷氨酰胺,以及蛋白质-谷氨酰胺谷氨酰胺酶(EC3.5.1.44;肽聚谷氨酰胺酶Ⅱ)特异于在羧基位置或在α-氨基和羧基位置上均有取代的谷氨酰胺。获自日本曲霉(Aspergillus japonicus),环状芽孢杆菌(Bacilluscirculans),Cryptococcns albidus和克洛德巴利酵母(Debaryomyceskloecheri)的肽聚谷氨酰胺酶已被报道。[Kikuchi M.和Sakaguchi K.;农业生物化学(Agr.Biol.Chem.)1973,37(4),719-724]。
已知肽聚谷氨酰胺酶用于改造食品蛋白质。蛋白质结构的修饰改善其功能性质且扩展了作为食品组分的用途。因此,US 5,082,672描述了用肽聚谷氨酰胺酶脱酰胺处理食品蛋白质的一种方法,通过此方法大豆蛋白质的溶解性、乳化作用、泡沫以及其它功能性质得以改善。然而,US 5,082,672也报道了由于其大的分子尺寸及/或独特构象使得肽聚谷氨酰胺酶对完整大豆蛋白的脱酰胺能力受到限制。因此,US 5,082,672指出底物的初始降解使用一种获自地衣芽孢杆菌的非特异性作用内/外肽酶复合物AlcalaseTM水解,且/或加热处理,并发现水解大豆蛋白的前热处理和后热处理产生最高的脱酰胺程度。
US 3,857,967描述了用获自环状芽孢杆菌的肽聚谷氨酰胺酶制备食品和饮料的方法。同样,为了获得最高的脱酰胺程度,US 3,857,967指出利用非特异性作用内/外肽酶的蛋白质底物的初始降解。
Mimouni等人[Minouni B.,Raymond J.Merle-Desnoyers A.M,AzanzaJ.L.和Ducastaing A.;谷物科学杂志(J.Cereal Science)1994,21,153-165]描述了用于改善小麦面筋的功能性质的联合酸脱酰胺和酶水解方法。更特别地,Mimouni等人描述了与使用非特异性作用内肽酶联合的酸脱酰胺方法。
已发现通过将蛋白质材料经过联合的脱酰胺作用处理及特异性作用的蛋白质水解酶作用后可获得具有极佳口味和功能的蛋白质水解物。
相应地,本发明提供了从蛋白质底物获得富含游离谷氨酸和/或肽结合的谷氨酸残基的水解物的一种方法,其包括以下步骤:底物经过脱酰胺处理;且将底物经过一种特异性作用蛋白水解酶作用。
在一优选的实施方案中,本发明提供了用于从蛋白质底物获得富含谷氨酸的水解物的一种方法,其包括以下步骤:将底物经过一种优选地切割谷氨酰-肽键的肽聚谷氨酰胺酶的作用且将底物经过一种蛋白水解酶作用,随后再经过一种或多种非特异性作用内和/或外肽酶作用。
本发明涉及一种获得富含游离谷氨酸和/或肽结合的谷氨酸残基的水解物的方法,其包括以下步骤:
(ⅰ)将该底物经过一种脱酰胺过程;且
(ⅱ)将该底物经过一种特异性作用蛋白水解酶的作用。
此两步骤,(ⅰ)和(ⅱ),可以同时完成,或步骤(ⅱ)也可在步骤(ⅰ)后进行。
通过本发明的方法可以获得口味和功能极佳的蛋白水解物。特别的,可获得极佳口味的蛋白水解物是由于谷氨酸(Glu),无论是游离的还是肽结合的,均对蛋白水解物的口味和风味起重要的作用。然而,通过本发明的方法,也可以获得功能改善了的蛋白质水解物,特别是在改善的溶解性、改善的乳化性质、增加的水解程度以及改善的泡沫性质方面。脱酰胺过程
通过放出氨进行的酰胺(谷氨酰胺或天冬酰胺)到荷电的酸(谷氨酸或天冬氨酸)的转化称为脱酰胺。脱酰胺可以是非酶法或酶法脱酰胺过程。
在优选的实施方案中,脱酰胺按一种酶法脱酰胺过程来进行。特别是通过将底物经过一种转谷氨酰胺酶或肽聚谷氨酰胺酶作用的方法进行酶法处理。转谷氨酰胺酶
在优选的实施方案中脱酰胺是用酶法脱酰胺过程来进行的,其中的底物经过一种转谷氨酰胺酶的作用。谷氨酰胺酶可以是任何方便的来源,包括那些来源自哺乳动物的,见如:JP 1050382和JP5023182,包括活化的第ⅩⅢ因子,见如:WO 93/15234;那些来源自鱼类的,见如:EP 555,649以及那些来源于微生物的,见如:EP 379,606,WO 96/06931和WO 96/22366。
在另一特殊的实施方案中,转谷氨酰胺酶来源于一种卵菌纲(Oomycete),包括疫霉属(Phytophthora)菌株,特别是恶疫霉(Phytophthora cactorum),腐霉属(Pythium)菌株,特别是畸雌腐霉(Pythium irregulare),腐霉属(Pythium sp.),间型腐霉(Pythiumintermedium),终极腐霉(Pythium ultimum)和周雄腐霉(Pythiumperiilum)(或缠器腐霉(P.periplocum))。
在另一特殊的实施方案中,转谷氨酰胺酶是细菌来源的,且优选的源自芽孢杆菌菌株,特别是枯草芽孢杆菌,链轮丝菌属(Streptoverticillium)菌株,特别是茂原链轮丝菌(Streptoverticilliummobaraensis),灰肉色链轮丝菌(Streptoverticillium griseocarneum),肉桂链轮丝菌(Streptoverticillium cinnamoneum)以及链霉菌属菌株,特别是利迪链霉菌。
转谷氨酰胺酶应以有效量加入到蛋白质底物中。转谷氨酰胺酶可以通常用于脱酰胺过程的量加入。目前考虑转谷氨酰胺酶的有效量在从约0.01到约5%(w/w)的相对于底物量的酶制剂的范围内,优选的在从约0.1到约1%(w/w)的相对于底物量的酶制剂的范围内。
酶法处理(孵育)可在任何方便的不使酶制剂失活的温度下进行,优选地在从约20℃到约70℃的范围内。
根据现有的实践,酶制剂可以被适当的失活,这或是通过升高培养混合物的温度到使酶失活的温度,如到高于约70℃,或是类似地通过降低培养混合物的pH到使酶失活的值,如低于约4.0。肽聚谷氨酰胺酶
在另一优选的实施方案中,脱酰胺是用一种酶法脱酰胺过程进行,其中的底物经过一种肽聚谷氨酰胺酶的作用。
在更特别的实施方案中,用于本发明的方法的肽聚谷氨酰胺酶可以是肽聚谷氨酰胺酶Ⅰ(肽酰谷氨酰胺酶;EC3.5.1.43)或肽聚谷氨酰胺酶Ⅱ(蛋白质谷氨酰胺谷氨酰胺酶;EC3.5.1.44)或其混合物。肽聚谷氨酰胺酶可以源自曲霉属的菌株,特别是日本曲霉,芽孢杆菌属的菌株,特别是环状芽孢杆菌,隐球酵母属(Cryptococcus)菌株,特别是Cryptococcus albidus,或德巴利酵母属(Debaryomyces)菌株,特别是克洛德巴利酵母菌。
肽聚谷氨酰胺酶应以有效量加入到蛋白质底物中。肽聚谷氨酰胺酶可以通常用于脱酰胺过程的量加入。目前考虑的肽聚谷氨酰胺酶的有效量范围以从约0.01到约100.000PG酶单位(PGase Unit)每100g底物的量,特别是从约0.1到约10.000PG酶单位每100g底物的量加入到底物中。
酶法处理(孵育)可在任何方便的不使酶制剂失活的温度下进行,优选地在从约20℃到约70℃的范围内。
根据现有的实践,酶制剂可以被适当的失活,这或是通过升高培养混合物的温度到使酶失活的温度,如到高于约70℃,或是类似地通过降低培养混合物的pH到使酶失活的值,如低于约4.0。特异性作用蛋白水解酶
本发明的方法包括将底物经过特异性作用蛋白水解酶作用的步骤。更特别的,特异性作用蛋白水解酶可以是一种特异性作用内肽酶或特异性作用外肽酶。特异性作用蛋白水解酶可以是任何可得的来源的。内肽酶
在优选的实施方案中,特异性作用蛋白水解酶是一种内肽酶,如:
(ⅰ)一种谷氨酰内肽酶(EC3.4.21.19);
(ⅱ)一种赖氨酰内肽酶(EC3.4.21.50);
(ⅲ)一种亮氨酰内肽酶(EC3.4.21.57);
(ⅳ)一种甘氨酰内肽酶(EC3.4.22.25);
(ⅴ)一种脯氨酰内肽酶(EC3.4.21.26);
(ⅵ)胰蛋白酶(EC3.4.21.4)或一种胰蛋白酶样内肽酶;或
(ⅶ)一种肽酰天冬氨酸金属内肽酶(EC3.4.24.33)。
项(ⅰ)的谷氨酰内肽酶(EC3.4.21.19)是一种蛋白水解酶,优选地切割谷氨酰肽键,其优选的源自芽孢杆菌属的菌株,特别是地衣芽孢杆菌和枯草芽孢杆菌,葡萄球菌属的菌株,特别是金黄色葡萄球菌,链霉菌属的菌株,特别是热普通链霉菌和灰色链霉菌,或是放线菌属的菌株。
项(ⅱ)的赖氨酰内肽酶(EC3.4.21.50)可以是优选的源自无色杆菌属(Achromobacter)的菌株,特别是水解无色杆菌(Achromobacterlyticus),溶杆菌属(Lysobacter)的菌株,特别是产酶溶杆菌(Lysobacterenzymogenes),或是假单胞菌属(Pseudomonas)的菌株,特别是铜绿假单胞菌(Pseudomonas aeruginosa)。
项(ⅲ)的亮氨酰内肽酶(EC3.4.21.57)可以是植物来源的。
项(ⅳ)的甘氨酰内肽酶(EC3.4.22.25)可优选的源自番木瓜属植物(Carica papaya)。
项(ⅴ)的脯氨酰内肽酶(EC3.4.21.26)可优选的源自黄杆菌属(Flavobacterium)的菌株,或可以是植物来源的。
项(ⅵ)的胰蛋白酶样内肽酶可优选的源自镰孢霉属(Fusarium)的菌株,特别是尖镰孢霉(Fusarium oxysporum)如WO89/06270或WO94/25583所述。
项(ⅶ)的肽酰天冬氨酸金属内肽酶(EC3.4.24.33)可优选的源自假单胞菌属的菌株,特别是霉实假单胞菌(Pseudomonas fragi)。外肽酶
在另一优选的实施方案中,特异性作用蛋白水解酶是一种外肽酶,其可以作用于肽的任一末端,即可以是氨肽酶或可以是羧肽酶。
在一优选的实施方案中,特异性作用蛋白水解酶是一种氨肽酶,如:
(ⅰ)一种亮氨酰氨肽酶(EC3.4.11.1);或
(ⅱ)一种三肽氨肽酶(EC3.4.11.4)。
在另一优选的实施方案中,特异性作用蛋白水解酶是一种羧肽酶,如:
(ⅰ)一种脯氨酸羧肽酶(EC3.4.16.2);
(ⅱ)一种羧肽酶A(EC3.4.17.1);
(ⅲ)一种羧肽酶B(EC3.4.17.2);
(ⅳ)一种羧肽酶C(EC3.4.16.5);
(ⅴ)一种羧肽酶D(EC3.4.16.6);
(ⅵ)一种赖氨酸(精氨酸)羧肽酶(EC3.4.17.3);
(ⅶ)一种甘氨酸羧肽酶(EC3.4.17.4);
(ⅷ)一种丙氨酸羧肽酶(EC3.4.17.6);
(ⅸ)一种谷氨酸羧肽酶(EC 3.4.17.11);
(ⅹ)一种肽酰二肽酶A(EC3.4.15.1);或
(ⅹⅰ)一种肽酰二肽酶(EC3.4.15.5)。
特异性作用的内或外肽酶应当以有效量加入蛋白质底物。蛋白水解酶可以通常用于蛋白质水解过程中的量加入,目前考虑内肽酶的有效量是以范围从约0.05到约15CPU/100g底物的量,特别是以范围从约0.1到约5CPU/100g底物,或以范围从约0.001到约0.5AU/100g底物,特别是范围从约0.01到约0.1AU/100底物的量。
酶法处理(孵育)可在任何方便的不使酶制剂失活的温度下进行,优选地在从约20℃到约70℃的范围内。
根据现有的实践,酶制剂可以被适当的失活,这或是通过升高培养混合物的温度到使酶失活的温度,如到高于约70℃,或是类似地通过降低培养混合物的pH到使酶失活的值,如低于约4.0。非特异性作用蛋白水解酶
在一更特别的实施方案中,本发明的方法包括附加的步骤:
(ⅲ)将底物经过一种或多种非特异性作用内和/或外肽酶作用的处理。
在此步骤中,底物经过传统水解步骤。这一附加步骤可以与步骤(ⅰ)和(ⅱ)同时完成,或其在步骤(ⅰ)和(ⅱ)之后完成。特别的,底物可以是步骤(ⅰ)和(ⅱ)的反应混合物。
在一优选的实施方案中,非特异性作用内和/或外肽酶源自曲霉属菌株,特别是黑曲霉菌株,米曲霉菌株,或是酱油曲霉菌株(Asperglliussoyae),或是芽孢杆菌属菌株,特别是解淀粉芽孢杆菌菌株(Bacillusamyloliquefaciens),迟缓芽孢杆菌(Bacillus lentus)菌株,地衣芽孢杆菌(Bacillus licheniformis)菌株或枯草芽孢杆菌菌株。
非特异性作用内和/或外肽酶以范围从约0.05到约15CPU/100g底物的量,特别是以范围从约0.1到约5CPU/100g底物的量加入到底物中。
酶法处理(孵育)可在任何方便的不使酶制剂失活的温度下进行,优选地在从约20℃到约70℃的范围内。
根据现有的实践,酶制剂可以被适当的失活,这或是通过升高培养混合物的温度到使酶失活的温度,如到高于约70℃,或是类似地通过降低培养混合物的pH到使酶失活的值,如低于约4.0。蛋白质底物
用本发明方法处理的蛋白质底物可含有完整的蛋白质,或其可含有预水解的蛋白质(肽),或其可为上述物质的混合物。同样,蛋白质底物可以是蔬菜或动物来源的。
优选的蛋白质底物为蔬菜来源的,特别是大豆蛋白质,谷物蛋白质,如:小麦面筋,玉米面筋,大麦,黑麦,燕麦,水稻,玉米醇溶蛋白,棉籽蛋白质,油菜籽蛋白质,花生,苜蓿蛋白质,豌豆蛋白质,蚕豆蛋白质,芝麻蛋白质或向日葵。
动物来源的蛋白质底物可特别是乳清蛋白,酪蛋白、肉类蛋白、鱼类蛋白、红血细胞,卵蛋白、明胶或乳白蛋白。工业应用
利用本发明的方法获得的富含游离谷氨酸和/或肽结合的谷氨酸残基的水解物可以用于不同的工业应用,特别是需要功能性蛋白质掺入的地方。
因此,另一方面,本发明提供了一种用本发明的方法获得的富含游离谷氨酸和/或肽结合的谷氨酸残基的水解物的食品。
在另一方面,本发明提供了一种用本发明的方法获得的富含游离谷氨酸和/或肽结合的谷氨酸残基的水解物的动物食品添加剂。酶活性的鉴定肽聚谷氨酰胺酶活性
肽聚谷氨酰胺酶活性可以根据Cedrangoro等人的方法来测定[Cedrangoro等人,Enzvmologia 1965,29,143],且如US 3,857,967中所述。
根据此方法,用1N的NaOH调节0.5ml的此酶样品至pH6.5,装满一小容器中。按Cedrangoro等人的方法,容器中加入1ml pH10.8的硼酸盐缓冲液,释放出的氨由5N硫酸吸收,利用Nessler试剂使混合物形成颜色,在420mγ处测量所形成的颜色的光吸收值。
在这些条件下能产生1γmol/分钟氨的酶的量为一个PG酶单位。
此外,肽聚谷氨酰胺酶活性可以按如实施例2(下面)所述的方法测定蛋白水解活性:酪蛋白蛋白酶单位(CPU)
利用酪蛋白为底物可测定蛋白水解活性。一酪蛋白蛋白酶单位(CPU)定义为在标准条件(即:25℃及pH9.5下温育30分钟)下每分钟放出1mM初级氨基基团(用与丝氨酸标准比较确定)的酶的量。
更详细描述此分析方法的文件AF 228/1只要需要可从NovoNordisk A/S丹麦获得,此处引用该文件作为参考。蛋白水解活性:Anson单位(AU)
也可以用变性血红蛋白为底物测定蛋白水解活性。在用于蛋白水解活性测定的Anson-血红蛋白方法中,消化变性血红蛋白,未消化的血红蛋白用三氯乙酸(TCA)沉淀。TCA溶解产物的量用苯酚试剂测定,其能与酪氨酸或色氨酸产生蓝色。
一个Anson单位(AU)定义为:在标准条件下(即:25℃,pH7.5以及反应时间为10分钟)以初始速度消化血红蛋白以使每分钟释放出的TCA溶解产物的量与苯酚试剂所产生的颜色和千分之一等量的酪氨酸相同。
更详细描述此分析方法的文件AF4/5只要需要就可从Novo NordiskA/S丹麦获得,此处引用作为参考。蛋白水解活性:亮氨酸氨肽酶单位(LAPU)
也可用亮氨酰-对-硝基苯胺为底物测定蛋白水解活性。一旦水解,对-硝基苯胺释放出来使溶液变黄。利用对-硝基苯胺为发色团测定活性,用光度计在405nm测产生的黄色。
一亮氨酸氨肽酶单位(LAPU)是在下列条件下每分钟分解1μ(微)M底物的酶的量:26mM亮氨酰-对-硝基苯胺为底物,0.1MTris缓冲液(pH8.0),40℃,反应时间10分钟。
实施例
本发明用下列实施例进一步阐明,其不是意在以任何方式限制本发明如权利要求所述的范围。实施例1
蛋白质溶解性提高和通过脱酰胺释放谷氨酸水解
小麦面筋(WG)获自Cargill(JOB 5141),脱酰胺小麦面筋(DWG)获自StaPro Consultancy B.V.,Lemdijk 32,9422 TH Smilde,NL。通过用89g水与11g面筋混合制成8%蛋白质悬浮液。用NaOH调节pH至6.5。谷氨酸/天冬氨酰特异性蛋白酶(SP446)(可如WO91/13554中所述获得),或赖氨酸/精氨酸特异性蛋白酶(SP387)(可如WO 89/06270中所述获得)加到悬浮液中。对SP446剂量为0.01AU/克蛋白质,对SP387为0.006AU/克蛋白质。以20LAPU/克蛋白质的量向一些水解物中加入FLavourzymeTM(一种可获自Novo Nordisk A/S,丹麦的非特异性作用蛋白酶制剂,含有内和外肽酶活性,且通过米曲霉发酵获得)。在无需进一步调整pH50℃条件下进行水解18小时。通过85℃加热15分钟使酶失活。调pH到5,离心水解物。测定在上清中的蛋白质以及游离谷氨酸的含量。蛋白质脱酰胺
用Kjeldahl分析法测定蛋白质,使用kjeldahl因子为6.25。谷氨酸的测定
通过使用适于谷氨酸测定的Boehringer Mannheim试剂盒(Cat.No.139092)测定游离谷氨酸的含量。将此方法改造成适于微滴定板上的应用。
结果
水解物 | 蛋白质溶解度%WG DWG | 谷氨酸含量mg/lWG DWG |
SP446 | 18 54 | 0 0 |
SP387 | 35 44 | 0 0 |
SPU446+FlavourzymeTM | 34 87 | 1000 1000 |
当比较小麦面筋(WG)和脱酰胺小麦面筋(DWG)时,结果表明脱酰胺作用增加了面筋对特异性蛋白酶的敏感性,从而使更多的蛋白质变得可溶。通过加入FlavourzymeTM到特异性蛋白酶上,由于脱酰胺作用谷氨酸的释放增加一倍。实施例2酶法脱酰胺作用和谷氨酸释放环状芽孢杆菌的发酵
ATCC21590菌株的环状芽孢杆菌培养物在容量为400ml内含200ml培养基的摇瓶中生长,培养基含有1%聚胨;0.5%乳糖;0.025%MgSO4·7H2O;0.005%FeSO4·7H2O;0.025%KH2PO4和17%Na2HPO4·12H2O,pH调至7.2。发酵在30℃下以搅拌速度270转/分进行20小时。以4000转/分离心收集细胞于1升摇瓶中。冷冻细胞。从环状芽孢杆菌中纯化肽聚谷氨酰胺酶
所有步骤均在室温下操作。
冷冻的环状芽孢杆菌细胞融化后悬浮于裂解缓冲液(50mMTris/HCl;25%(w/v)蔗糖;1mM EDTA,pH8.0)直到获得均质悬浮液,每升裂解缓冲液100g湿细胞。LysozymeTM(Fluka 62971,10mg/ml)和DNAseⅠ(Sigma DN-25,10mg/ml)溶于裂解缓冲液。加入100mlLysozymeTM溶液,10ml1.0M MgCl2和1ml DNAseⅠ溶液到每升细胞悬浮液中。使酶作用1小时。
用Seitz深层过滤板过滤悬浮液,滤液移至10mM KH2PO4/NaOH,pH8.0(缓冲液A)的Sephadex G25柱(Pharmacia)上。酶溶液加到用缓冲液A平衡过的SOURCE Q柱(Pharmacia)上并以缓冲液A中的线性NaCl梯度(0→500mM)洗脱。如下述分析柱流分的肽聚谷氨酰胺酶Ⅱ活性,且合并有活性的流分。所合并的流分的280nm吸光值为1.78,因此估计其蛋白质含量为1.8mg/ml。
从SDS-PAGE凝胶推断肽聚谷氨酰胺酶Ⅱ合并液中蛋白质的纯度约为25%。因此制剂中含有约0.5mg/ml的纯肽聚谷氨酰胺酶Ⅱ。肽聚谷氨酰胺酶鉴定方法
通过使用适于氨测定的Boehringer-Mannheim试剂盒(Cat.No.1112732)测量在N-叔丁氧羰基(N-t-BOC-Gln-Pro;SIGMA No.B-4406)γ-羧酰胺的水解过程中形成的氨确定酶活性。在此试剂盒中,氨是通过测定由谷氨酸脱氢酶消耗的NADH来确定的,不加入N-t-BOC-Gln-Pro的空白也进行测量以抵消其它消耗NADH的酶的影响。小麦面筋水解
200mg小麦面筋蛋白质加入到9ml沸水中,冷却后,调节pH到7.0。再加入如上述的250μl肽聚谷氨酰胺酶Ⅱ制剂(PEP)。以0.04AU/g蛋白质的量加入来自实施例1的谷氨酸/天冬氨酸特异性蛋白酶(SP446),且以20LAPU/克蛋白质的量加入来自实施例1的FlavourzymeTM。
在50℃不调节pH条件下水解进行18小时。不加入肽聚谷氨酰胺酶的空白也同样处理。离心水解物且如实施例1所述测定谷氨酸。
用如下述方式测定小麦面筋蛋白质的水解(DH)程度。水解程度(DH)的测定
通过上清液与(OPA)(邻苯二甲醛,Sigma)的反应测定蛋白质的水解程度,如Adler-Nissen[Adler-Nissen J.;食物蛋白质的酶水解(Enzymic Hydrolysis of Food Proteins);Elsevier应用科学出版社,1986]所述。对OPA试剂,160mg OPA溶于4ml乙醇,并移入容积200ml含有7.62g十水合四硼酸二钠和200mg十二烷基硫酸钠以及176mg二硫苏糖醇的烧瓶中,用水加至烧瓶的标记。试剂是光敏感的。
25μl适当稀释的上清液与200μl OPA试剂在微滴定板的孔中混合,且在25℃下反应恰好2分钟。在微孔板读数仪上测定340nm吸光值,且与95mM L-丝氨酸标准溶液在减去空白值(与OPA试剂反应的水)后的吸光值比较。为找出真实的水解程度,在上清液中测得的丝氨酸当量用由Adler-Nissen用于三硝基苯磺酸方法的推荐因子修正[Adler-Nissen J.;农业和食品化学(Agricultural and Food Chemistry),1979,27(6),1256],其与所述的OPA方法有相同的反应。水解程度在水解混合物中总蛋白质的量的基础上计算出(不以溶解的蛋白质为基础)。结果
水解 | DH% | 谷氨酸mg/l |
不加PEP | 40 | 131 |
加PEP | 43 | 171 |
如表所示,使用肽聚谷氨酰胺酶制剂的水解增加水解的程度以及谷氨酸的释放。
Claims (33)
1.一种从蛋白质底物获得一种富含游离谷氨酸和/或肽结合的谷氨酸残基的水解物的方法,包括以下步骤:
(ⅰ)将该底物经过一种脱酰胺过程;且
(ⅱ)将该底物经过一种特异性作用蛋白水解酶的作用。
2.如权利要求1的方法,其中的脱酰胺过程按一种非酶法脱酰胺进行。
3.如权利要求1的方法,其中的脱酰胺过程按一种酶法脱酰胺进行。
4.如权利要求3的方法,其中的酶法脱酰胺过程通过将底物经过一种转谷氨酰胺酶的作用来进行。
5.如权利要求5的方法,其中的转谷氨酰胺酶源自卵菌纲,包括疫霉属的菌株,特别是恶疫霉,腐霉属的菌株,特别是畸雌腐霉,腐霉属,间型腐霉,终极腐霉和周雄腐霉(或缠器腐霉)。
6.如权利要求5的方法,其中的转谷氨酰胺酶是细菌来源的,且优选源自芽孢杆菌属的菌株,特别是枯草芽孢杆菌,链轮丝菌属的菌株,特别是茂原链轮丝菌,灰肉色链轮丝菌和肉桂链轮丝菌,以及链霉菌属的菌株,特别是利迪链霉菌。
7.如权利要求3的方法,其中的酶法脱酰胺过程通过将底物经过一种肽聚谷氨酰胺酶的作用来进行。
8.如权利要求7的方法,其中的肽聚谷氨酰胺酶是肽聚谷氨酰胺酶Ⅰ(肽酰谷氨酰胺酶;EC3.5.1.43)或肽聚谷氨酰胺酶Ⅱ(蛋白质谷氨酰胺谷氨酰胺酶;EC3.5.1.44)或其混合物。
9.如权利要求7或8的方法,其中的肽聚谷氨酰胺酶源自曲霉属的菌株,特别是日本曲霉,芽孢杆菌属的菌株,特别是环状芽孢杆菌,隐球酵母属的菌株,特别是Cryptococcus albidus,或德巴利酵母属的菌株,特别是克洛德巴利酵母菌。
10.如权利要求7-9中任一项的方法,其中的肽聚谷氨酰胺酶以范围从约0.01到约100.000的PG酶单位每100g底物的量,特别是以约0.1到约10.000PG酶单位每100g底物的量加入到底物中。
11.如权利要求1-10中任一项的方法,其中特异性作用蛋白水解酶是一种内肽酶,如:
(ⅰ)一种谷氨酰内肽酶(EC3.4.21.19);
(ⅱ)一种赖氨酰内肽酶(EC3.4.21.50);
(ⅲ)一种亮氨酰内肽酶(EC3.4.21.57);
(ⅳ)一种甘氨酰内肽酶(EC3.4.22.25);
(ⅴ)一种脯氨酰内肽酶(EC3.4.21.26);
(ⅵ)胰蛋白酶(EC3.4.21.4)或一种胰蛋白酶样内肽酶;或
(ⅶ)一种肽酰天冬氨酸金属内肽酶(EC 3.4.24.33)。
12.如权利要求11的方法,其中(ⅰ)的谷氨酰内肽酶源自芽孢杆菌属的菌株,特别是地衣芽孢杆菌和枯草芽孢杆菌,葡萄球菌属的菌株,特别是金黄色葡萄球菌,链霉菌属的菌株,特别是热普通链霉菌和灰色链霉菌,或是放线菌属的菌株。
13.如权利要求11的方法,其中(ⅱ)的赖氨酰内肽酶是源自无色杆菌属的菌株,特别是水解无色杆菌,溶杆菌属的菌株,特别是产酶溶杆菌,或是假单胞菌属的菌株,特别是铜绿假单胞菌。
14.如权利要求11的方法,其中(ⅳ)的甘氨酰内肽酶源自番木瓜属植物(Carica papaya)。
15.如权利要求11的方法,其中(ⅴ)的脯氨酰内肽酶源自黄杆菌属的菌株。
16.如权利要求11的方法,其中(ⅵ)的胰蛋白酶样内肽酶源自镰孢霉属的菌株,特别是尖镰孢霉。
17.如权利要求11的方法,其中(ⅶ)的肽酰天冬氨酸金属内肽酶(EC3.4.24.33)源自假单胞菌属的菌株,特别是霉实假单胞菌。
18.如权利要求1-16项中任一项的方法,其中的特异性作用蛋白水解酶是一种外肽酶。
19.如权利要求17的方法,其中的外肽酶是一种氨肽酶,如:
(ⅰ)一种亮氨酰氨肽酶(EC 3.4.11.1);或
(ⅱ)一种三肽氨肽酶(EC 3.4.11.4)。
20.如权利要求17的方法,其中的外肽酶是一种羧肽酶,如:
(ⅰ)一种脯氨酸羧肽酶(EC3.4.16.2);
(ⅱ)一种羧肽酶A(EC3.4.17.1);
(ⅲ)一种羧肽酶B(EC3.4.17.2);
(ⅳ)一种羧肽酶C(EC3.4.16.5);
(ⅴ)一种羧肽酶D(EC3.4.16.6);
(ⅵ)一种赖氨酸(精氨酸)羧肽酶(EC3.4.17.3);
(ⅶ)一种甘氨酸羧肽酶(EC3.4.17.4);
(ⅷ)一种丙氨酸羧肽酶(EC3.4.17.6);
(ⅸ)一种谷氨酸羧肽酶(EC3.4.17.11);
(ⅹ)一种肽酰二肽酶A(EC3.4.15.1);或
(ⅹⅰ)一种肽酰二肽酶(EC3.4.15.5)。
21.如权利要求7-10项中任一项的方法,其从蛋白质底物获得富含谷氨酸的水解物,此方法包括如下步骤:
(ⅰ)将此底物经过肽聚谷氨酰胺酶作用;且
(ⅱ)将此底物再经优选地切割谷氨酰肽键的蛋白水解酶作用(即:一种谷氨酰内肽酶;EC3.4.21.19)。
22.如权利要求1-21中任一项的方法,其中特异性作用蛋白水解酶以范围从约0.05到约15CPU/100g底物的量,特别是在范围从约0.1到约5CPU/100g底物的量加入到底物中。
23.如权利要求1-22中任一项的方法,其中权利要求1的步骤(ⅰ)和步骤(ⅱ)是同时完成的。
24.如权利要求1-22中任一项的方法,其中权利要求1的步骤(ⅱ)是在权利要求1的步骤(ⅰ)之后完成的。
25.如权利要求1-24中任一项的方法,其方法包括附加步骤:
(ⅲ)将底物经过一种或多种非特异性作用内和/或外肽酶的作用。
26.如权利要求25的方法,其中步骤(ⅰ),(ⅱ)和(ⅲ)是同时完成的。
27.如权利要求25的方法,其中步骤(ⅲ)是在步骤(ⅰ)-(ⅱ)之后完成的。
28.如权利要求25-27中任一项的方法,其中步骤(ⅲ)的非特异性作用内和/或外肽酶源自曲霉属菌株,特别是黑曲霉菌株,米曲霉菌株,或是酱油曲霉菌株,或是芽孢杆菌属的菌株,特别是解淀粉芽孢杆菌菌株,迟缓芽孢杆菌菌株,地衣芽孢杆菌菌株或枯草芽孢杆菌菌株。
29.如权利要求25-28中任一项的方法,其中的非特异性作用内和/或外肽酶以范围从约0.05到约15CPU/100g底物的量,特别是以范围从约0.1到约5CPU/100g底物,或以范围从约0.001到约0.5AU/100g底物,特别是范围从约0.01到约0.1AU/100g底物的量加入到底物中。
30.如权利要求1-29中任一项的方法,其中的蛋白质底物为蔬菜来源的,特别是大豆蛋白质,谷物蛋白质,如:小麦面筋,玉米面筋,大麦,黑麦,燕麦,水稻,玉米醇溶蛋白,棉籽蛋白质,油菜籽蛋白质,花生,苜蓿蛋白质,豌豆蛋白质,蚕豆蛋白质,芝麻蛋白质或向日葵。
31.如权利要求1-29中任一项的方法,其中的蛋白质底物为动物来源的,特别是乳清蛋白,酪蛋白,肉类蛋白,鱼类蛋白,红血细胞,卵蛋白,明胶或乳白蛋白。
32.一种含有按权利要求1-31中任一项的方法获得的富含游离谷氨酸和/或肽结合的谷氨酸残基的水解物的食品。
33.一种含有按权利要求1-31中任一项的方法获得的富含游离谷氨酸和/或肽结合的谷氨酸残基的水解物的动物食品添加剂。
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Families Citing this family (26)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
WO1998051804A1 (en) * | 1997-05-16 | 1998-11-19 | Novo Nordisk Biotech, Inc. | Polypeptides having aminopeptidase activity and nucleic acids encoding same |
AU7381098A (en) * | 1997-05-16 | 1998-12-08 | Novo Nordisk Biotech, Inc. | Polypeptides having prolyl pipeptidyl aminopeptidase activity and nucleic acids encoding same |
US6251651B1 (en) | 1998-06-04 | 2001-06-26 | Amano Pharmaceutical Co., Ltd. | Protein-deamidating enzyme, gene encoding the same, production process therefor, and use thereof |
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Family Cites Families (4)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
US3857967A (en) * | 1971-05-10 | 1974-12-31 | Kikkoman Shoyu Co Ltd | Preparation of food and beverages with peptidoglutaminase |
US5082672A (en) * | 1989-06-21 | 1992-01-21 | The United States Of American As Represented By The Secretary Of Agriculture | Enzymatic deamidation of food proteins for improved food use |
JP2814300B2 (ja) * | 1989-09-01 | 1998-10-22 | プロテイン テクノロジーズ インターナシヨナル インコーポレーテツド | 酵素変性蛋白と方法 |
ES2077220T3 (es) * | 1990-03-09 | 1995-11-16 | Novo Nordisk As | Hidrolizados de proteinas. |
-
1997
- 1997-05-20 WO PCT/DK1997/000230 patent/WO1997043910A1/en active IP Right Grant
- 1997-05-20 JP JP09541414A patent/JP2000515003A/ja active Pending
- 1997-05-20 BR BR9709006A patent/BR9709006A/pt active Search and Examination
- 1997-05-20 CN CN97194838A patent/CN1099839C/zh not_active Expired - Fee Related
- 1997-05-20 EP EP97921649A patent/EP0969736B1/en not_active Expired - Lifetime
- 1997-05-20 DE DE69736247T patent/DE69736247T2/de not_active Expired - Fee Related
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2001
- 2001-05-31 CN CN01119457A patent/CN1326687A/zh active Pending
Cited By (6)
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CN103354718A (zh) * | 2010-12-06 | 2013-10-16 | 卡吉尔公司 | 用于液化谷物蛋白的方法 |
CN103354718B (zh) * | 2010-12-06 | 2016-03-23 | 卡吉尔公司 | 用于液化谷物蛋白的方法 |
CN104982891A (zh) * | 2015-06-11 | 2015-10-21 | 广东厨邦食品有限公司 | 一种酱油原油的制备方法 |
CN105248835A (zh) * | 2015-09-02 | 2016-01-20 | 广东美味鲜调味食品有限公司 | 玉米蛋白水解物及其制备方法与在纯粮酿造食醋中的应用 |
CN105248835B (zh) * | 2015-09-02 | 2019-09-20 | 广东美味鲜调味食品有限公司 | 玉米蛋白水解物及其制备方法与在纯粮酿造食醋中的应用 |
FR3057180A1 (fr) * | 2016-12-12 | 2018-04-13 | Constellium Issoire | Procede d'amelioration du mouillage d'une surface d'un substrat solide par un metal liquide |
Also Published As
Publication number | Publication date |
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AU2765097A (en) | 1997-12-09 |
CN1099839C (zh) | 2003-01-29 |
BR9709006A (pt) | 1999-08-03 |
DE69736247T2 (de) | 2007-05-24 |
AU722915B2 (en) | 2000-08-17 |
JP2000515003A (ja) | 2000-11-14 |
EP0969736A1 (en) | 2000-01-12 |
CN1326687A (zh) | 2001-12-19 |
WO1997043910A1 (en) | 1997-11-27 |
DE69736247D1 (de) | 2006-08-10 |
EP0969736B1 (en) | 2006-06-28 |
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