CN107683334A - 具有改进的去污力的包含至少一种漆酶的洗涤剂 - Google Patents
具有改进的去污力的包含至少一种漆酶的洗涤剂 Download PDFInfo
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- CN107683334A CN107683334A CN201680032343.3A CN201680032343A CN107683334A CN 107683334 A CN107683334 A CN 107683334A CN 201680032343 A CN201680032343 A CN 201680032343A CN 107683334 A CN107683334 A CN 107683334A
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- laccase
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Abstract
本发明涉及特定的漆酶在洗涤织物中的用途,并涉及包含所述漆酶的洗涤剂。
Description
本发明涉及特定的漆酶在清洗织物过程中的用途,以及包含所述漆酶的洗涤剂。
漆酶(EC 1.10.3.2)是存在于许多植物、真菌和微生物中的含铜的“蓝色”酶。漆酶属于氧化还原酶。催化活性中心含有四个铜离子,它们的光谱性质可能不同。“蓝色”1型铜参与底物氧化;一个2型和两个3型铜离子形成三核簇,其与氧键合并且还原成水。漆酶也称为对苯二酚氧化酶。除了苯二酚之外,漆酶氧化许多其它底物,例如甲氧基取代的苯酚和二胺。就其底物而言,漆酶意外地是非特异性的。由于其广泛的底物特异性和氧化酚类化合物的能力,漆酶引起了人们对其工业应用的极大兴趣。许多有希望应用漆酶的领域包括例如木材工业中纤维板的脱木质化和粘合,纺织工业中为物质着色和净化染料废水以及用于生物传感器。
在介质即中间分子的帮助下,漆酶还可以氧化其原本不能氧化的底物。介质通常是可被漆酶氧化的“小分子化合物”。然后氧化的介质又氧化实际的底物。
首个漆酶在1883年发现于日本漆树(Rhus vernicifera)。漆酶可见于许多植物中,如桃、番茄、芒果和马铃薯;漆酶还已知在一些昆虫中。然而,最常使用的漆酶来自真菌,例如来自伞菌属(Agaricus)、曲霉属(Aspergillus)、下皮黑孔菌属(Cerrena)、弯孢属(Curvularia)、镰孢属(Fusarium)、香菇属(Lentinius)、单链孢属(Monocillium)、毁丝霉属(Myceliophtora)、脉孢菌属(Neurospora)、青霉菌属(Penicillium)、平革菌属(Phanerochaete)、射脉菌属(Phlebia)、侧耳属(Pleurotus)、柄孢壳菌属(Podospora)、裂褶菌属(Schizophyllum)、孢子丝菌属(Sporotrichum)、壳多孢属(Stagonospora)和栓菌属(Trametes)。
本质上,漆酶的功能尤其包括参与木质纤维素的分解、细胞壁的生物合成、水果和蔬菜的褐变以及防止微生物对植物的攻击等。
通常难以有效地从脏衣物或脏物中去除有色污渍/杂质。去除强着色污渍和杂质、即来自水果和/或蔬菜的污渍和杂质是特别需求的。这些污渍和杂质含有基于类胡萝卜素化合物的有色色素,例如a-、b-和g-胡萝卜素和番茄红素、卟啉类如叶绿素、以及类黄酮色素和染料组分。基于天然类黄酮的最后一组染料组分包含基于天竺葵素、花青色素、花翠素及其甲酯和花黄素的强着色花青苷染料和色素。这些化合物是水果中的大部分橙色、红色、紫色和蓝色的来源,并富含于所有浆果、樱桃、红醋栗和黑醋栗、葡萄柚、百香果、橙子、柠檬、苹果、梨、石榴、红甘蓝、甜菜根中,且富含于花中。花青色素的衍生物存在于多达80%的有色叶、多达70%的水果和多达50%的花中。这种杂质的特定实例包括来自茶、咖啡、调味料如咖喱和红辣椒、橙子、番茄、香蕉、茶、芒果、西兰花、胡萝卜、甜菜根、菠菜和草的杂质。圆珠笔墨水也被认为是很难去除的污渍。这些强着色的类黄酮和类胡萝卜素染料通常是具有共轭双键体系的多环和杂环化合物。这种化学结构通常是化合物的清晰颜色的原因。
配制液体洗涤剂和清洁剂时,除去有色污渍和杂质的所述困难特别严重,因为这些制剂通常不含任何漂白剂。
令人惊讶地发现,迄今为止未知的来自凤尾菇(Pleurotus pulmonarius)的漆酶的SEQ.ID NO.1或与其足够相似的漆酶(对于序列相同性而言)特别适用于液体洗涤剂或清洁剂,并且改进去污力。
因此,本发明的第一主题是一种漆酶,其包含在其总长度上与SEQ ID NO.1所示氨基酸序列至少70%、渐增优选至少75%、80%、81%、82%、83%、84%、85%、86%、87%、88%、89%、90%、90.5%、91%、91.5%、92%、92.5%、93%、93.5%、94%、94.5%、95%、95.5%、96%、96.5%、97%、97.5%、98%、98.5%和99%相同的氨基酸序列。
本发明的另一主题是一种漆酶,其由在其总长度上与SEQ ID NO.2所示核酸序列至少70%、渐增优选至少75%、80%、81%、82%、83%、84%、85%、86%、87%、88%、89%、90%、90.5%、91%、91.5%、92%、92.5%、93%、93.5%、94%、94.5%、95%、95.5%、96%、96.5%、97%、97.5%、98%、98.5%和99%相同的核酸序列编码。
本发明的另一主题是一种洗涤剂,其包含至少一种本发明的漆酶,即包含在其总长度上与SEQ ID NO.1所示氨基酸序列至少70%、渐增优选至少75%、80%、81%、82%、83%、84%、85%、86%、87%、88%、89%、90%、90.5%、91%、91.5%、92%、92.5%、93%、93.5%、94%、94.5%、95%、95.5%、96%、96.5%、97%、97.5%、98%、98.5%和99%相同的氨基酸序列的漆酶,或由在其总长度上与SEQ ID NO.2所示核酸序列至少70%、渐增优选至少75%、80%、81%、82%、83%、84%、85%、86%、87%、88%、89%、90%、90.5%、91%、91.5%、92%、92.5%、93%、93.5%、94%、94.5%、95%、95.5%、96%、96.5%、97%、97.5%、98%、98.5%和99%相同的核酸序列编码的漆酶。
有利地,本发明的制剂具有改进的去污力,特别是在去除有色污渍和杂质时。特别优选的是,本发明的制剂适用于去除含花青苷的杂质。
SEQ ID NO.1是来自担子菌纲(Basidiomycete)凤尾菇DSMZ 5331(DSMZ:德国微生物菌种保藏中心)的漆酶的序列。
SEQ ID NO.2是编码SEQ ID NO.1的漆酶的cDNA,即没有内含子。
此外,令人惊奇地发现,以下漆酶也特别适用于液体洗涤剂或清洁剂,并且改进去污力:
a)来自担子菌纲佛罗里达平菇(Pleurotus ostreatus var.florida)的漆酶[SEQ.ID NO.3]或与其足够相似的漆酶(相对于序列相同性而言);
b)来自香菇真菌(埃杜拉香菇(Lentinula edodes))的至今未知的漆酶[SEQ.IDNO.5]或与其足够相似的漆酶(相对于序列相同性而言);
c)来自栓菌属种(Tsp,DSMZ 11309)(DSMZ:德国微生物菌种保藏中心)的至今未知的漆酶[SEQ.ID NO.7]或与其足够相似的漆酶(相对于序列相同性而言)。
因此,本发明的另一主题是一种漆酶,其包含在其总长度上与SEQ ID NO.3、SEQID NO.5和/或SEQ ID NO.7所示氨基酸序列至少70%、渐增优选至少75%、80%、81%、82%、83%、84%、85%、86%、87%、88%、89%、90%、90.5%、91%、91.5%、92%、92.5%、93%、93.5%、94%、94.5%、95%、95.5%、96%、96.5%、97%、97.5%、98%、98.5%和99%相同的氨基酸序列。
因此,本发明的另外主题是洗涤剂,其包含至少一种包含下述氨基酸序列的漆酶,所述氨基酸序列在其总长度上与SEQ ID NO.3、SEQ ID NO.5和/或SEQ ID NO.7所示氨基酸序列至少70%、渐增优选至少75%、80%、81%、82%、83%、84%、85%、86%、87%、88%、89%、90%、90.5%、91%、91.5%、92%、92.5%、93%、93.5%、94%、94.5%、95%、95.5%、96%、96.5%、97%、97.5%、98%、98.5%和99%相同。
有利地,本发明的制剂具有改进的去污力,特别是在去除有色污渍和杂质时。特别优选的是,本发明的制剂适用于去除含花青苷的杂质。
SEQ ID NO.3是来自佛罗里达平菇的漆酶的序列,其来自文献Palmieri,G.等(2003),Atypical laccaseisoenzymes from coppersupplemented Pleurotus ostreatuscultures,EnzymeMicrob.Technol.33(2-3),220-230。序列在该出版物的第227页。
SEQ ID NO.5是来自担子菌纲埃杜拉香菇(香菇)的漆酶的序列。
SEQ ID NO.7是来自栓菌属种(Tsp,DSMZ 11309)(DSMZ:德国微生物菌种保藏中心)的漆酶的序列。
本发明的另一主题是一种含有至少一种漆酶的洗涤剂,该漆酶由在其总长度上与SEQ ID NO.4、SEQ ID NO.6或SEQ ID NO.8所示核酸序列至少70%、渐增优选至少75%、80%、81%、82%、83%、84%、85%、86%、87%、88%、89%、90%、90.5%、91%、91.5%、92%、92.5%、93%、93.5%、94%、94.5%、95%、95.5%、96%、96.5%、97%、97.5%、98%、98.5%和99%相同的核酸序列编码。
SEQ ID NO.4是编码SEQ ID NO.3的漆酶的cDNA,即没有内含子。
SEQ ID NO.6是编码SEQ ID NO.5的漆酶的cDNA,即没有内含子。
SEQ ID NO.8是编码SEQ ID NO.7的漆酶的cDNA,即没有内含子。
通过序列比较确定核酸或氨基酸序列的相同性。该序列比较基于现有技术中确立并常规使用的BLAST算法(参见例如Altschul,S.F.,Gish,W.,Miller,W.,Myers,E.W.&Lipman,DJ.(1990)“Basic local alignment search tool.”J.Mol.Biol.215:403410,和Altschul,Stephan F.,Thomas L.Madden,Alejandro A.Schaffer,Jinghui Zhang,HhengZhang,Webb Miller和David J.Lipman(1997):“Gapped BLAST and PSI-BLAST:a newgeneration of protein database search programs”;Nucleic Acids Res.,25,pp.3389-3402)并采取在核酸或氨基酸序列中相互指定的相似核苷酸序列或氨基酸序列的形式。各个位置的列表分配称为比对。现有技术中可用的另一种算法是FASTA算法。使用计算机程序生成序列比较(比对),特别是多序列比较。经常使用的是例如Clustal系列(参见例如Chenna等(2003):Multiple sequence alignment with the Clustal series ofprograms.Nucleic Acid Research 31,3497-3500)、T-Coffee(参见例如Notredame等(2000):T-Coffee:Anovel method for multiple sequencealignmentments.J.Mol.Biol.302,205-217)或基于这些程序或算法的程序。在本专利申请中,使用具有给定标准参数的计算机程序VectorSuite 10.3(InvitrogenCorporation,1600Faraday Avenue,Carlsbad,California,USA)产生所有序列比较(比对),用于序列比较的AlignX模块基于ClustalW。
此种比较也使得其可以对比较序列的相似性进行评价。其通常以相同性百分比表示,即相同位置处或在比对中对应于另一位置处的相同核苷酸或氨基酸残基的比例。在氨基酸序列中,同源性的更广义阐述的术语涉及保守氨基酸交换,因此是指具有相似化学活性的氨基酸,因为这些氨基酸在蛋白质内发挥主要相似的化学活性。因此,比较序列之间的相似性也可以表示为同源性百分比或相似性百分比。相同性和/或同源性的细节可以通过整个多肽或基因或仅通过个别区域发现。因此,不同核酸序列或氨基酸序列的同源或相同区域由序列中的相似性定义。这些区域通常具有相同的功能。它们可以是小的并且仅包含几个核苷酸或氨基酸。通常,这样的小区域对蛋白质的整体活性发挥必要的功能。因此,将序列相似性仅与单个的、可能的小区域相关联可能是有利的。然而,除非另有说明,在本发明中,相同性或同源性细节是指分别指出的核酸序列或氨基酸序列的全长。
本发明的漆酶和可用于本发明的洗涤剂的漆酶可以从植物、真菌并优选细菌获得,特别是从杆菌和放线菌获得。包含SEQ ID NO.1的漆酶可以从凤尾菇获得。包含SEQ IDNO.3的漆酶可以从佛罗里达平菇获得。包含SEQ ID NO.5的漆酶可以从埃杜拉香菇获得。包含SEQ ID NO.7的漆酶可以从栓菌属种获得。
然而,漆酶的天然产量通常非常低。因此,通过在外源生产宿主中表达漆酶基因来增加产量是有利的。
为此目的,通常使用包含编码本发明的漆酶的核酸的载体。
这些可以是DNA分子或RNA分子。它们可以作为单链存在,作为与该单链互补的单链或双链存在。特别地,对于DNA分子,两个互补链的序列分别在所有三个可能的阅读框中考虑。此外,应认为不同密码子即碱基三联体可以编码相同氨基酸,结果可以从几种不同的核酸编码特定的氨基酸序列。本领域技术人员能够明确地确定这些核酸序列,因为尽管遗传密码已经简并,但可以将各个密码子分配给所定义的氨基酸。因此,从氨基酸序列开始,本领域技术人员可以没有问题地确定编码该氨基酸序列的核酸。此外,在核酸中,一个或多个密码子可以被同义密码子替代。该方面特别涉及本发明的酶的异源表达。以这种方式,每个生物体例如生产菌株的宿主细胞都具有特定的密码子使用。密码子使用应理解为是指由相应生物体将遗传密码翻译成氨基酸。如果核酸上的密码子在生物体内面临相对少数量的加载(belandenen)tRNA分子,则蛋白质生物合成可能缺乏。虽然编码相同氨基酸,但是这导致密码子在生物体中的翻译效率低于编码相同氨基酸的同义密码子。由于存在更多数量的用于同义密码子的tRNA分子,因此可以在生物体中更有效地翻译。
使用本领域技术人员现今通常已知的方法,例如化学合成或结合分子生物学和/或蛋白质化学标准方法的聚合酶链式反应(PCR),可以使用已知的DNA和/或氨基酸序列产生直至完整基因的相应核酸。这种方法已知于例如Sambrook,J.,Fritsch,E.F.andManiatis,T.2001.Molecular cloning:a laboratory manual,冷泉港实验室出版社,第三版。
在本发明的意义内,可理解载体是指由核酸组成的元件,该元件含有编码本发明的漆酶的作为表征核酸区域的核酸。他们能够将这些区域作为物种或细胞系中历经几代或多次细胞分裂的稳定的遗传元件。载体是特殊的质粒,因此是环状遗传元件,特别是在细菌中使用时。在本发明的框架内,将编码本发明的漆酶的核酸克隆到载体中。载体包括例如来源于细菌质粒、病毒或噬菌体的载体,或主要为合成载体或具有最多多种来源的元件的质粒。随着其他遗传元件分别可用,载体能够将其本身建立为在相关宿主细胞的几代中的稳定单位。它们可以作为单独的单位存在于染色体外,或整合到染色体或染色体DNA中。
表达载体包含使其能够在含有它们的宿主细胞优选微生物、特别优选细菌中复制并表达其中所含的核酸的核酸序列。该表达特别是通过调控转录的启动子来影响。原则上,表达可以通过原本位于待表达核酸之前的天然启动子进行,也可以通过表达载体上提供的宿主细胞的启动子或通过另一个生物体或另一个宿主细胞的经修饰或完全不同的启动子进行。在本申请中,使至少一种启动子用于表达编码本发明的漆酶的核酸,并用于其表达。还可以调控表达载体,例如通过改变培养条件或含有其的宿主细胞达到特定细胞密度、或通过添加特定物质(特别是基因表达的激活剂)。这种物质的实例是用作细菌乳糖操纵子(lac-操纵子)的激活剂的半乳糖衍生物异丙基β-D-1-硫代吡喃半乳糖苷(IPTG)。与表达载体不同,所含的核酸不在克隆载体中表达。
优选地,将编码本发明的漆酶的核酸或含有这种核酸的载体转化进入微生物,该微生物然后用作宿主细胞。或者,可以将各个组分即编码本发明的漆酶的核酸的核酸部分或片段引入宿主细胞,使得所得宿主细胞含有一种此类核酸或一种此类载体。如果宿主细胞已经含有此类核酸或此类载体的一种或多种组分或几种组分,则这种方法特别合适,并且随后可相应地补充其它组分。转化细胞的方法在现有技术中已经建立,并且是本领域技术人员充分已知的。原则上,所有细胞即原核细胞或真核细胞都适合作为宿主细胞。优选可以有利地进行遗传操作的那些宿主细胞,例如其影响核酸或载体的转化及其稳定建立,例如单细胞真菌或细菌。此外,优选的宿主细胞特征在于良好的微生物学和生物技术操纵行为。例如,这涉及容易培养、高生长速度、对发酵培养基的需求小以及对于外源蛋白质的良好生产和分泌速率。本发明优选的宿主细胞将(转基因)表达的蛋白质分泌到宿主细胞周围的培养基中。此外,漆酶可以通过产生其的细胞在生产后进行修饰,例如通过连接糖分子、甲酰化、酰胺化等进行修饰。这种翻译后修饰可以在功能上影响漆酶。
其活性因遗传调控元件而可被调控的此类宿主细胞特别适用于生产本发明的漆酶,所述遗传调控元件例如可以在载体上可获得,但也可以在这些细胞中提前存在。例如,通过监测添加作为激活剂的化学化合物,通过改变培养条件或达到特定细胞密度时,可以激励这些化合物表达。这使得本发明的蛋白质的经济生产成为可能。此类联系的一个例子是IPTG,如前所述。
优选的宿主细胞是原核细胞或细菌细胞。细菌特征在于世代时间短,对培养条件的要求小。因此,建立了具有成本优势的培养方法或生产方法。此外,发酵技术领域的技术人员将具有丰富的细菌经验。由于多种多样的原因,如在个别情况中实验建立的营养成分来源、产品形成率和所需时间等,革兰氏阴性菌或革兰氏阳性菌可适用于特殊生产。
对于革兰氏阴性菌如大肠杆菌等,多种蛋白质分泌在周质空间中,从而分泌到包围细胞的膜之间的区室中。这对于特殊应用是有利的。此外,还可以开发革兰氏阴性菌,使得它们不仅在周质空间中排出表达的蛋白质,而且在细菌周围的培养基中排出表达的蛋白质。另一方面,革兰氏阳性菌例如杆菌或放线菌或放线菌的其他代表没有外膜,结果是分泌的蛋白质立即排放在细菌周围的培养基(通常是营养培养基)中,从而可以纯化表达的蛋白质。它们可以直接从培养基中分离或进一步处理。此外,革兰氏阳性菌与大多数产生技术上重要的酶的原始生物相关或相同,并且它们本身形成大多数可比较的酶,结果是它们具有相似的密码子使用,并且其蛋白质合成器官自然相应对应。
所述宿主细胞可以根据其对培养条件的要求而改变,或者具有不同的或额外的选择标记,或表达不同的或额外的蛋白质。特别地,这些宿主细胞也可以是那些转基因表达几种蛋白质或酶的宿主细胞。
本发明原则上可以应用于所有微生物,特别是所有可发酵的微生物,并且导致能够通过使用这种微生物产生本发明的漆酶。
用于获得本发明的漆酶的特别优选的宿主细胞是细菌,特别是那些选自以下属的细菌:埃希氏菌属、克雷伯菌属、芽孢杆菌属、葡萄球菌属、棒杆菌属、节杆菌属、链霉菌属、寡养单胞菌属和假单胞菌属,特别是选自:大肠杆菌(Escherichia coli)、植生克雷伯菌(Klebsiella planticola)、地衣芽孢杆菌(Bacillus licheniformis)、迟缓芽孢杆菌(Bacillus lentus)、解淀粉芽孢杆菌(Bacillus amyloliquefaciens)、枯草芽孢杆菌(Bacillus subtilis)、嗜碱芽孢杆菌(Bacillus alcalophilus)、球芽孢杆菌(Bacillusalcalophilus)、吉氏芽孢杆菌(Bacillus gibsonii)、克劳氏芽孢杆菌(Bacillusclausii)、耐盐芽孢杆菌(Bacillus halodurans)、短小芽孢杆菌(Bacillus pumilus)、肉葡萄球菌(Staphylococcus carnosus)、谷氨酸棒杆菌(Corynebacterium glutamicum)、氧化节杆菌(Arthrobacter oxidans)、变铅青链霉菌(Streptomyces lividans)、天蓝色链霉菌(Streptomyces coelicolor)和嗜麦芽寡养单胞菌(Stenotrophomonas maltophilia)。
然而,宿主细胞也可以是真核细胞,其特征在于其具有细胞核。与原核细胞相比,真核细胞可以以翻译后方式修饰形成的蛋白质。这些细胞的实例是真菌,例如放线菌或酵母菌,如酵母菌属或克鲁维酵母属。例如,如果与蛋白质合成相结合,这种系统使得可能使蛋白质经历特定修饰,这可能是特别有利的。执行真核系统特别是与蛋白质合成相结合的修饰包括例如键合低分子化合物如膜锚或寡糖。这种寡糖修饰在例如用于降低所表达的蛋白质的变应原性时可能是期望的。此外,与由这些细胞天然形成的酶(例如纤维素酶或脂肪酶)的共表达可能是有利的。此外,例如嗜热真菌表达系统可能特别适用于表达耐热蛋白质或变体。
所述宿主细胞例如在不连续或连续系统中常规培养和发酵。在第一种情况中,将宿主细胞接种在合适的营养培养基并通过实验确定在一段时间后从培养基中收获的产物。连续发酵特征在于实现动态平衡,其中细胞在相当长的一段时间内部分地死亡,但也可以重新长出,并且形成的蛋白质可以同时从培养基中移走。
发酵方法本身是现有技术中已知的,并且代表实际的工业生产步骤,随后通常是通过合适的纯化方法纯化所得产品(例如本发明的漆酶)。
特别考虑到特征在于通过补料策略进行发酵的发酵方法。通过连续培养而消耗的培养基成分由此得到补充。因此,可以在细胞密度以及细胞团或干质量和/或特别是目的漆酶的活性方面实现相当大的增长。此外,发酵还可以发展使不期望的代谢产物通过加入缓冲液或各自合适的平衡离子过滤掉或中和。
所生产的漆酶可以从发酵培养基中收获。这种发酵方法相对于从宿主细胞分离漆酶、即从细胞团(干质量)的产品制备是优选的,但需要提供合适的宿主细胞或一种或多种合适的分泌标记或机制和/或转运系统,以使宿主细胞将漆酶分泌到发酵培养基中。或者无需分泌,漆酶即可从宿主细胞中分离,即从细胞团中将其纯化出,例如使用常规的酶化学方法,例如盐沉淀、超滤、离子交换色谱和疏水相互作用色谱。纯化可通过SDS聚丙烯酰胺凝胶电泳进行监测。可以测定纯化酶在不同温度和pH值的酶活性;类似地,可以测定分子量和等电点。
令人惊奇的是,已经发现本发明的漆酶,特别是包含SEQ ID NO.1的漆酶以及包含SEQ ID NO.3、SEQ ID NO.5和SEQ ID NO.7的漆酶可改善去污力,特别是改善液体洗涤剂制剂的去污力,并且不像已知的其它漆酶那样引起不期望的变暗和由此带来的污渍(特别是茶渍和咖啡渍)加剧。
本发明的洗涤剂中漆酶的浓度相对于活性蛋白为0.001~0.15wt%,优选为0.005~0.06wt%。
本发明的洗涤剂可以优选在5℃~95℃的温度范围内使用,优选20℃~60℃,特别优选30℃~40℃。
本发明的洗涤剂可以含有另外的调节剂,以便以更高的效率氧化待除去的染料。本发明适合的调节剂是例如Tempo(2,2,6,6-四甲基-1-哌啶基氧)、HBT(1-羟基苯并三唑)、ABTS(2,2'-连氮双-3-乙基苯并噻唑-6-磺酸酯)、NHA(N-羟基-乙酰苯胺)、2,5-二甲苯胺、乙醇、铜、4-甲基邻苯二酚、N-羟基邻苯二甲酰亚胺、没食子酸、单宁酸、槲皮素、丁香酸、愈创木酚、二甲氧基苄醇、苯酚、紫脲酸(异亚硝基巴比土酸)、酚红、溴酚蓝、纤维素、对香豆素酸、路易波士(Rooibos)、邻甲酚、二氯靛酚、羟基苯并三唑、环己酰亚胺或香草醛。
本发明的另一主题是本发明的漆酶用于改善洗涤剂的去污力的用途,特别是用于去除有色污渍和杂质,特别优选用于除去含有花青苷的杂质。
本发明的另一主题是在含有表面活性剂的水溶液中洗涤织物的方法,其特征在于,使用含有表面活性剂的水溶液含有至少一种本发明的漆酶。
该方法以其最简单的形式实现,即将需要清洁的织物与所述水液接触,其中可以使用常规的洗衣机或者可以手工洗衣。根据本发明,优选如同使用常规家用洗衣机程序的情况,使用洗液强力冲洗来实施所述方法。
除了适于除去所述污渍和杂质的漆酶之外,洗涤剂还可以含有通常与这种组分相容的成分。因此,例如,其可以另外含有一种或多种颜色转移抑制剂,其优选的量为0.1wt%~2wt%,特别是0.2wt%~1wt%,在优选的实施方案中,其选自乙烯基吡咯烷酮、乙烯基咪唑、乙烯基吡啶-N-氧化物的聚合物或其共聚物。可以使用分子量为15 000g/mol~50000g/mol的聚乙烯吡咯烷酮和分子量更高的聚乙烯吡咯烷酮(例如高达1 000 000g/mol以上,特别是1500000g mol~4000000g/mol)、N-乙烯基咪唑/N-乙烯基吡咯烷酮共聚物、聚乙烯基噁唑烷酮、基于乙烯基单体和羧酸酰胺的共聚物、含有吡咯烷酮基团的聚酯和聚酰胺、接枝聚酰胺胺和聚乙烯亚胺、聚胺-N-氧化物聚合物和聚乙烯醇。还可以使用包含过氧化物酶和过氧化氢或在水中提供过氧化氢的物质的酶系统。在这种情况下,优选添加过氧化物酶的调节剂化合物,例如乙酰丁香酮、苯酚衍生物或吩噻嗪或吩噁嗪,其中还可以另外使用上述聚合物色素转移抑制剂活性成分。聚乙烯吡咯烷酮的平均摩尔质量优选在10 000g/mol~60 000g/mol的范围内,特别是在25 000g/mol~50 000g/mol的范围内。在共聚物中,优选由摩尔比为5:1~1:1的乙烯基吡咯烷酮和乙烯基咪唑制成的共聚物,其平均摩尔质量在5 000g/mol~50 000g/mol范围内,特别是10 000g/mol~20 000g/mol。
洗涤剂可以特别是颗粒粉末固体、再稠化颗粒形式、粒状形式、均一溶液或悬浮液存在,除了本发明使用的漆酶以外,其原则上还可含有在此种制剂中已知和常见的所有成分。本发明的制剂可以特别包含助洗剂(Buildersubstanzen)、表面活性表面活性剂、基于有机和/或无机过氧化合物的漂白剂、漂白活化剂、水溶性有机溶剂、酶、螯合剂、电解质、pH调节剂和进一步的助剂,例如荧光增白剂、发灰抑制剂(Vergrauungsinhibitoren)、泡沫调节剂以及染料和香味剂。
所述制剂优选包含一种表面活性剂或几种表面活性剂,其中特别地考虑阴离子表面活性剂、非离子表面活性剂及其混合物,还有阳离子、两性离子和两性表面活性剂。
合适的非离子表面活性剂特别是烷基糖苷和烷基糖苷或直链或支链醇的乙氧基化和/或丙氧基化产物,每个在烷基部分中具有12~18个碳原子与3~20、优选4~10个烷基醚基团。此外,可以使用对于烷基部分而言对应所述长链醇衍生物的N-烷基胺、邻位二醇、脂肪酸酯和脂肪酸酰胺以及烷基残基中具有5~12个碳原子的烷基酚的相应乙氧基化和/或丙氧基化产物。
优选烷氧基化的、有利地为乙氧基化的,特别是具有优选8~18个碳原子且每摩尔醇平均1~12摩尔环氧乙烷(EO)的伯醇用作非离子表面活性剂,其中醇残基可以是线性或优选在2位有甲基支链,或者可以在混合物中含有线性和甲基支链残基,像羰基合成醇残基中常规存在的一样。然而,特别是具有由带有12~18个碳原子的天然来源的醇制成(例如由椰子、棕榈、牛油或油醇制成)的线性残基和每摩尔醇平均2~8个EO的醇乙氧基化物是优选的。优选的乙氧基化醇包括例如具有3个EO或4个EO的C12-C14醇,具有7个EO的C9-C11醇,具有3个EO、5个EO、7个EO或8个EO的C13-C15醇,具有3个EO、5个EO或7个EO的C12-C18醇及其混合物,例如具有3个EO的C12-C14醇与具有7个EO的C12-C18醇的混合物。所指出的乙氧基化度表示统计平均数,其对于特殊产品可以是整数或是分数。优选的醇乙氧基化物具有集中的同系物分布(窄范围乙氧基化物,NRE)。除了这些非离子表面活性剂之外,还可以使用具有超过12个EO的脂肪醇。其实例是具有14个EO、16个EO、20个EO、25个EO、30个EO或40个EO的(牛油)脂肪醇。使用常规的极低发泡化合物,特别是在用于机器方法的制剂中。它们优选包括C12-C18烷基聚乙二醇聚丙二醇醚,每个在分子中具有至多8摩尔环氧乙烷和环氧丙烷单元。然而,也可以使用其它已知的低发泡非离子表面活性剂,例如C12-C18烷基聚乙二醇聚丁二醇醚(每个在分子中各自具有至多8摩尔环氧乙烷和环氧丁烷单元)以及末端封端的烷基聚亚烷基二醇混合醚。含羟基的烷氧基化醇(所谓的羟基混合醚)()也是特别优选的。可以使用的非离子表面活性剂还包括通式为RO(G)x的烷基糖苷,其中R表示主要的直链或甲基支链脂肪族残基,特别是2位甲基支链,具有8~22、优选12~18个碳原子,G表示具有5或6个碳原子的葡萄糖单元,优选为葡萄糖。表示单糖苷和寡糖苷分布的寡聚化程度x是任何数值,其也可以采取分数值作为分析测定的变量,为1~10;优选x为1.2~1.4。还合适的是下式的多羟基脂肪酸酰胺:
其中R1CO表示具有6~22个碳原子的脂肪族酰基残基,R2表示氢、具有1~4个碳原子的烷基或羟烷基残基,[Z]表示具有3~10个碳原子和3~10个羟基的直链或支链多羟基烷基残基。
优选地,多羟基脂肪酸酰胺衍生自具有5或6个碳原子的还原糖,特别是葡萄糖。多羟基脂肪酸酰胺的基团还包括下式的化合物:
其中,R3表示具有7~12个碳原子的直链或支链烷基或烯基残基,R4表示直链、支链或环状亚烷基残基或具有2~8个碳原子的亚芳基残基,R5表示具有1~8个碳原子的直链、支链或环状烷基残基或芳基残基或烷氧基残基,其中C1-C4烷基或苯基残基是优选的,并且[Z]表示直链多羟基烷基残基(其烷基链被至少两个羟基取代)、或烷氧基化、优选乙氧基化或丙氧基化的该残基的衍生物。还优选通过糖(例如葡萄糖、果糖、麦芽糖、乳糖、半乳糖、甘露糖或木糖)的还原胺化来获得[Z]。通过在作为催化剂的醇盐存在下与脂肪酸甲酯反应,可以将N-烷氧基取代或N-芳氧基取代的化合物转化为所需的多羟基脂肪酸酰胺。作为单一非离子表面活性剂使用或与其它非离子表面活性剂组合使用(特别是与烷氧基化脂肪醇和/或烷基糖苷一起使用)的另一类优选使用的非离子表面活性剂是经烷氧基化、优选乙氧基化或乙氧基化和丙氧基化的脂肪酸烷基酯,其优选在烷基链中具有1~4个碳原子,特别是脂肪酸甲酯。此外,胺氧化物类型的非离子表面活性剂,例如N-椰油烷基-N,N-二甲基氧化胺和N-牛脂烷基-N,N-二羟基乙基氧化胺和脂肪酸烷醇酰胺可以是合适的。这些非离子表面活性剂的量优选不超过乙氧基化脂肪醇的量,特别是不超过其一半。还考虑所谓的双子(Gemini)表面活性剂作为其它表面活性剂。通常,这些表面活性剂是指每分子具有两个亲水基团的那些化合物。这些基团通常通过所谓的“间隔区”彼此分离。该间隔区通常是碳链,其应该足够长以使亲水基团具有足够的距离,使得它们可以彼此独立地起作用。这种表面活性剂通常特征在于非常小的临界胶束浓度和强烈降低水表面张力的能力。在特殊情况下,表述双子表面活性剂不仅理解为“二聚”,而且也表示相应的“三聚”表面活性剂。合适的双子表面活性剂是例如硫酸化羟基混合醚或二聚醇-双-和三聚醇-三-硫酸盐和-醚硫酸盐。末端封端的二聚和三聚混合醚特征特别在于它们的双官能性和多官能性。因此,所述末端封端的表面活性剂具有良好的交联性能和低发泡性,使其特别适用于机洗或清洗方法。然而,也可以使用双子多羟基脂肪酸酰胺或聚多羟基脂肪酸酰胺。
合适的阴离子表面活性剂特别是皂和那些含有硫酸盐或磺酸盐基团的。作为磺酸盐型表面活性剂,可考虑的是优选C9-C13烷基苯磺酸盐、烯烃磺酸盐,即烯烃与羟基烷烃磺酸盐和二磺酸盐的混合物,例如为通过用气态三氧化硫磺化以及随后磺化产物的碱水解或酸水解得到的具有末端或内部双键的C12-C18单烯烃。此外,例如通过磺基氯化或磺基氧化以及随后的水解或中和而从C12-C18烷烃获得的烷烃磺酸盐是合适的。此外,α-磺基脂肪酸的酯(磺酸酯)是合适的,例如氢化椰子脂肪酸、棕榈仁脂肪酸或牛油脂肪酸的α-磺化甲酯,其通过在脂肪酸分子中具有8~20个碳原子的植物和/或动物来源的脂肪酸甲酯的α-磺化和随后中和为水溶性单盐而制得。优选地,其是氢化椰油脂肪酸、棕榈脂肪酸、棕榈仁脂肪酸或牛油脂肪酸的α-磺化酯,其中还可以少量存在不饱和脂肪酸(例如油酸)的磺化产物,其量优选不高于约2~3wt%。特别地,优选具有在酯基中具有不超过4个C原子的烷基链的α-磺基脂肪酸烷基酯,例如甲酯、乙酯、丙酯和丁酯。特别有利的是,使用α-磺基脂肪酸的甲基酯(MES),也可以使用其皂化的二盐。其它合适的阴离子表面活性剂是代表单酯、二酯和三酯及其混合物的磺基脂肪酸甘油酯,如得自通过单甘油与1~3mol脂肪酸的酯化生产期间或在甘油三酯与0.3~2mol甘油的酯化期间。优选作为烷基(烯基)硫酸盐的是C12-C18脂肪酸醇(例如椰子油醇、牛油脂肪醇、月桂醇、肉豆蔻醇、十六烷醇或十八烷醇或C10-C20羰基合成醇)的硫酸半酯以及这些链长的仲醇的那些半酯的碱盐、特别是钠盐。还优选的是所述链长的烷基(烯基)硫酸盐,其含有以石油化学为基础产生的合成的直链烷基残基,其具有类似于基于脂肪化学原料的适当化合物的降解行为。从洗涤相关的技术方面来看,特别优选C12-C16烷基硫酸盐和C12-C15烷基硫酸盐以及C14-C15烷基硫酸盐。另外,可以作为ShellOil Company以商品名商业化商品获得的2,3-烷基硫酸盐也是合适的阴离子表面活性剂。还合适的是直链或支链C7-C21醇(例如具有平均3.5mol环氧乙烷(EO)的2-甲基支链的C9-C11醇或具有1~4个EO的C12-C18脂肪酸醇)的硫酸单酯,其中C7-C21醇用1~6mol环氧乙烷乙氧基化。优选的阴离子表面活性剂还包括烷基磺基琥珀酸的盐,其也称为磺基琥珀酸盐或磺基琥珀酸酯,并且其表示磺基琥珀酸与醇(优选脂肪醇,特别是乙氧基化脂肪醇)的单酯和/或二酯。优选的磺基琥珀酸盐含有C8-C18脂肪醇残基或其混合物。特别地,优选的磺基琥珀酸盐含有衍生自乙氧基化脂肪醇(其被认为本身代表非离子表面活性剂)的脂肪醇残基。反之,其脂肪醇残基衍生自具有集中的同系物分布的乙氧基化脂肪醇的磺基琥珀酸盐是特别优选的。同样地,还可以使用在烷基(烯基)链中优选具有8~18个碳原子的烷基(烯基)磺基琥珀酸或其盐。作为其它阴离子表面活性剂,考虑氨基酸的脂肪酸衍生物,例如N-甲基牛磺酸(Tauride)和/或N-甲基甘油(Sarkoside)。特别优选的是Sarkoside或肌氨酸盐,这里尤其是高级的且任选单或多不饱和的脂肪酸的肌氨酸盐,如油酰肌氨酸盐。特别地,考虑皂作为其他阴离子表面活性剂。特别地,饱和脂肪酸皂,例如月桂酸、肉豆蔻酸、棕榈酸、硬脂酸、氢化芥酸和山萮酸的盐、特别是来自天然脂肪酸(例如椰子,棕榈仁或牛油脂肪酸)的盐,或衍生的皂混合物是合适的。已知的烯基琥珀酸盐还可以与这些皂一起使用,或作为皂的替换物。
存在阴离子表面活性剂,包括其钠、钾或铵盐形式的皂以及有机碱的可溶性盐,例如单-、二-或三-乙醇胺。优选地,阴离子表面活性剂以其钠盐或钾盐的形式存在,特别是以钠盐的形式存在。表面活性剂通常以1wt%~50wt%、特别是5wt%~30wt%的量包含在洗涤剂中。
洗涤剂优选含有至少一种水溶性和/或水不溶性的有机和/或无机助洗剂。水溶性有机助洗剂物质包括聚羧酸,特别是柠檬酸和糖酸,单体和聚合的氨基聚羧酸,特别是甘氨酸二乙酸、甲基甘氨酸二乙酸、次氮基三乙酸、亚氨基二琥珀酸盐如乙二胺-N,N'-二琥珀酸和羟基亚氨基二琥珀酸盐、乙二胺四乙酸以及聚天冬氨酸、聚膦酸,特别是氨基三(亚甲基膦酸),乙二胺四(亚甲基膦酸)、赖氨酸四(亚甲基膦酸)和1-羟基乙烷-1,1-二膦酸,聚合羟基化合物如糊精以及聚合(多)羧酸、特别是可通过多糖、聚丙烯酸、甲基丙烯酸、马来酸及其混合聚合物的氧化获得的聚羧酸盐,其也可以含有少部分的聚合的不含羧酸官能的可聚合物质。在每种情况下相对于其游离盐而言,不饱和羧酸的均聚物的相对平均分子质量通常在5 000g/mol~200000g/mol,共聚物的相对平均分子质量在2 000g/mol~200 000g/mol,优选为50 000g/mol~120 000g/mol。特别优选的丙烯酸-马来酸共聚物的相对平均分子质量为50 000g/mol~100 000g/mol。如果也不太优选,合适的该类化合物是丙烯酸或甲基丙烯酸与乙烯基醚(如乙烯基甲基醚、乙烯基酯、乙烯、丙烯和苯乙烯)的共聚物,其中酸的比例为至少为50wt%。还可以使用含有作为单体的两种不饱和酸和/或其盐以及作为第三单体的乙烯醇和/或乙烯醇衍生物或碳水化合物的水溶性有机助洗剂三元共聚物。第一种酸单体或其盐衍生自单烯式不饱和C3-C8羧酸,优选衍生自C3-C4一元羧酸,特别是(甲基)丙烯酸。第二种酸单体或其盐可以是C4-C8二羧酸的衍生物,其中马来酸是特别优选的。在这种情况下,第三单体单元由乙烯醇和/或优选酯化的乙烯醇形成。特别地,优选乙烯醇衍生物,其代表短链羧酸(例如C1-C4羧酸)与乙烯醇的酯。优选的聚合物包含60wt%~95wt%、特别是70wt%~90wt%的(甲基)丙烯酸或(甲基)丙烯酸盐/酯(特别优选丙烯酸或丙烯酸盐/酯)和马来酸或马来酸盐/酯以及5wt%~40wt%、优选10wt%~30wt%的乙烯醇和/或乙酸乙烯酯。特别优选如下聚合物,其中(甲基)丙烯酸或(甲基)丙烯酸盐/酯与马来酸或马来酸盐/酯的重量比为1:1~4:1,优选为2:1~3:1,特别优选为2:1和2.5:1。数量和重量比均相对于酸而言。第二酸单体或其盐也可以是在2位取代为烷基残基(优选为C1-C4烷基残基)或优选衍生自苯或苯衍生物的芳族残基的烯丙基磺酸的衍生物。优选的三元共聚物包含40wt%~60wt%、特别是45~55wt%的(甲基)丙烯酸或(甲基)丙烯酸盐(特别优选丙烯酸或丙烯酸盐),10wt%~30wt%、优选15wt%~25wt%的甲代烯丙基磺酸或甲代烯丙基磺酸盐,以及作为第三单体的15wt%~40wt%、优选20wt%~40wt%的烃。该烃可以例如是单糖、二糖、寡糖或多糖,其中优选单糖、二糖或寡糖。蔗糖是特别优选的。推测通过使用第三单体将弱化点并入聚合物中,这些点对聚合物的良好生物降解性有贡献。这些三元共聚物的相对平均分子质量通常在1 000g/mol~200 000g/mol,优选在200g/mol~50 000g/mol。进一步优选的共聚物是具有丙烯醛和丙烯酸/丙烯酸盐或乙酸乙烯酯作为单体的那些共聚物。有机助洗剂可以以水溶液的形式使用,优选为30~50重量%的水溶液的形式,特别是用于生产液体制剂。所有上述酸通常以其水溶性盐、特别是其碱盐的形式使用。
必要时,可以包含此类有机助洗剂的量直至40wt%、特别是至多25wt%、优选1wt%~8wt%。接近所述上限的量优选用于糊剂或液体、特别是含水制剂中。
可考虑水溶性无机助洗剂材料,特别是多磷酸盐,优选三磷酸钠。作为水不溶性无机助洗剂材料,特别是结晶或无定形的无机助洗剂材料,可使用水分散性碱铝硅酸盐,其量不超过25wt%,优选3wt%~20wt%,并且量特别是5wt%~15wt%。洗涤剂品质中的结晶铝硅酸钠,特别是沸石A、沸石P和沸石MAP以及任选沸石X是优选的。接近所述上限的量优选用于固体颗粒剂。合适的铝硅酸盐特别地不具有粒径大于30μm的任何颗粒,并且由优选高达至少80wt%且尺寸小于10μm的颗粒构成。钙结合能力通常在100~200mg CaO/克的范围。
另外,或者对于所述水不溶性铝硅酸盐和碱碳酸盐来说,可以包含另外的水溶性无机助洗剂材料。除了多磷酸盐如三磷酸钠之外,这些材料包括特别是水溶性结晶和/或无定形碱硅酸盐助洗剂。这种水溶性无机助洗剂物质优选以1wt%~20wt%、特别是5wt%~15wt%的量包含在制剂中。可用作助洗剂材料的碱硅酸盐中碱氧化物与SiO2的摩尔比例优选小于0.95,特别是1:1.1~1:12,并且可以以无定形或结晶形式存在。优选的碱硅酸盐是硅酸钠,特别是无定形硅酸钠,Na2O:SiO2的摩尔比为1:2~1:2.8。作为结晶硅酸盐,其可以单独存在或存在于无定形硅酸盐的混合物中,优选使用通式为Na2SixO2x+1·y H2O的结晶层状硅酸盐,其中x(所谓的系数)为1.9~4的数,y为0~20的数,x的优选值为2、3或4。优选的结晶层状硅酸盐是其中所述通式中的x为2或3的那些。特别地,优选β-和δ-二硅酸钠(Na2Si2O5·y H2O)。此外,由上述通式的无定形碱硅酸盐制成的实际上无水的结晶碱硅酸盐可用于制剂中,其中x为1.9~2.1的数。在另一优选实施方案中,使用系数为2~3的结晶层状硅酸钠,其可由沙和苏打制得。在另一实施方案中使用系数为1.9~3.5的硅酸钠。在这种制剂的优选实施方案中,使用碱硅酸盐和碱碳酸盐的颗粒化合物,其例如以名称15市售可获得。
作为漂白剂,可考虑基于氯的那些化合物,例如特别是碱性次氯酸盐、二氯异氰脲酸及其盐,特别是还有基于过氧化物的那些。特别地,可考虑有机过酸或有机酸的过酸盐如苯二甲酰亚氨基己酸、过苯甲酸、单过氧苯二甲酸和二过十二烷酸及其盐作为合适的过氧化合物,例如单过氧苯二甲酸镁、过氧化氢、以及在使用条件下释放出过氧化氢的无机盐(如过硼酸盐、过碳酸盐和/或过硅酸盐,如过氧化氢包合物如H2O2尿素加合物)。也可以使用酶系统即氧化酶及其底物来生产过氧化氢。在使用固体过氧化合物的情况下,它们可以以粉末或颗粒的形式使用,其也可以以已知方式包封。特别优选的是,使用碱性过碳酸盐、碱性过硼酸盐一水合物或水溶液形式的过氧化氢(含有3wt%~10wt%过氧化氢)。如果洗涤剂含有过氧化合物,它们的存在量优选至多25wt%,特别是1wt%~20wt%,特别优选7wt%~20wt%。
特别地,在过水解条件下产生任选取代的过苯甲酸和/或具有1~12个碳原子、特别是2~4个碳原子的脂肪族过氧羧酸的化合物(单独或混合物)用作漂白活化化合物,其在过水解条件下提供过氧羧酸。具有特别是所述碳原子数的O-和/或N-酰基和/或任选具有取代的苯甲酰基的漂白活化剂是特别优选的。多种酰化亚烷基二胺是优选的,特别是四乙酰基乙二胺(TAED)、酰化甘脲特别是四乙酰基甘脲(TAGU)、酰化三嗪衍生物特别是1,5-二乙酰-2,4-二氧六氢-1,3,5-三嗪(DADHT)、N-酰基酰亚胺特别是N-壬酰琥珀酰亚胺(NOSI)、酰化苯酚磺酸盐或羧酸盐或其磺酸或羧酸、特别是壬酰基或异壬酰基或月桂酰氧基苯磺酸盐(NOBS或异-NOBS或LOBS)或癸酰氧基苯甲酸(DOBA)、其正式的碳酸衍生物如4-(2-癸酰基氧乙氧基羰基氧)-苯磺酸盐(DECOBS)、酰化多元醇特别是三乙酸甘油酯、乙二醇二乙酸酯和2,5-二乙酰氧基-2,5-二氢呋喃以及乙酰化山梨糖醇和甘露糖醇及其混合物(SORMAN)、酰化糖衍生物特别是五乙酰葡萄糖(PAG)、五乙酰果糖、四乙酰木糖和八乙酰乳糖、乙酰化且任选N-烷基化的葡萄糖胺和葡萄糖酸内酯和/或N-酰化内酰胺(例如N-苯甲酰基己内酰胺)。
除了在过水解条件下形成过氧羧酸的化合物之外,还可存在其它漂白活化化合物,例如腈类,其在过水解条件下形成过酰亚胺酸。这些化合物特别地包括下式的具有季铵化氢原子的氨基乙腈衍生物:
其中,R1表示-H、-CH3、C2-24烷基或烯基残基、具有选自-Cl、-Br、-OH、-NH2、-CN和-N(+)-CH2-CN中的至少一个取代基的取代的C1-24烷基或C2-24烯基残基、具有C1-24烷基的烷基或烯基芳基残基、或芳环上具有至少一个、优选两个、任选经取代的C1-24烷基和任选具有其他取代基的取代的烷基或烯基芳基残基,R2和R3彼此独立地选自-CH2-CN、-CH3、-CH2-CH3、-CH2-CH2-CH3、-CH(CH3)-CH3、-CH2-OH、-CH2-CH2-OH、-CH(OH)-CH3、-CH2-CH2-CH2-OH、-CH2-CH(OH)-CH3、-CH(OH)-CH2-CH3、-(CH2CH2-O)nH,其中n=1、2、3、4、5或6,R4和R5彼此独立地具有如前对R1、R2或R3所指出的含义,其中至少2个所述残基(特别是R2和R3)还包括氮原子和任选其他杂原子可以以闭环方式彼此连接,而后优选形成吗啉环,X是电荷平衡阴离子,优选选自苯磺酸根、甲苯磺酸根、异丙基苯磺酸根、C9-15烷基苯磺酸根、C1-20烷基硫酸根、C8-22羧酸甲酯磺酸根、硫酸根、硫酸氢根及其混合物。此外,可以使用转移氧的磺酰亚胺(Sulfonimine)和/或酰腙。
而且,可以存在漂白催化性过渡金属络合物。这些优选选自钴、铁、铜、钛、钒、锰和钌络合物。这些过渡金属络合物中可考虑无机和有机化合物作为配体,除了羧酸盐之外,那些化合物包括,特别是具有伯胺、仲胺和/或叔胺和/或醇官能团的化合物,例如吡啶、哒嗪、嘧啶、吡嗪、咪唑、吡唑、三唑、2,2'-双吡啶胺、三(2-吡啶基甲基)胺、1,4,7-三氮杂环壬烷、1,4,7-三甲基-1,4,7-三氮杂环壬烷、1,5,9-三甲基-1,5,9-三氮杂环十二烷、(双-((1-甲基咪唑-2-基)-甲基))-(2-吡啶基甲基)-胺、N,N'-(双-(1-甲基咪唑-2-基)-甲基)-乙二胺、N-双(2-苯并咪唑基甲基)-氨基乙醇、2,6-双-(双-(2-苯并咪唑基甲基)氨基甲基)-4-甲基苯酚、N,N,N',N'-四(2-苯并咪唑基甲基)-2-羟基-1,3-二氨基丙烷、2,6-双-(双-(2-吡啶基甲基)氨基甲基)-4-甲基苯酚、1,3-双-(双-(2-苯并咪唑基甲基)氨基甲基)-苯、山梨糖醇、甘露糖醇、赤藓糖醇、核糖醇、肌醇、乳糖,和任选取代的盐水、卟吩和紫菜碱。无机中性配体特别地包括氨和水。如果不是所有的过渡金属中心原子的配位中心都被中性配体占据,那么络合物还优选含有阴离子配体,尤其是单齿配体或双齿配体。这些配体特别地包括卤化物,如氟化物、氯化物、溴化物和碘化物以及(NO2)-基团,即硝基配体或亚硝基配体。(NO2)-基团可以以螯合方式与过渡金属键合,也可以不对称地或以η1-O方式桥接两个过渡金属原子。除了所述配体之外,过渡金属络合物可以具有其他的、通常结构简单的配体,特别是单价或多价阴离子配体。例如,可考虑硝酸根、乙酸根、三氟乙酸根、甲酸根、碳酸根、草酸根、高氯酸根和复合阴离子如六氟磷酸根。阴离子配体旨在确保过渡金属中心原子和配体体系之间的电荷均衡。此外,可以存在氧基配体、过氧基配体和亚氨基配体。特别地,这样的配体也可以是桥接的,结果产生多核络合物。在桥联双核络合物的情况下,络合物中的两个金属原子不必需相同。此外,可以使用其中两个过渡金属中心原子具有不同氧化数的双核络合物。如果阴离子配体丢失,或阴离子配体的存在不会导致络合物中的电荷均衡,则在根据本发明使用的过渡金属络合物化合物中,存在中和阳离子过渡金属络合物的阴离子平衡离子。这些阴离子平衡离子特别地包括硝酸根、氢氧根、六氟磷酸根、硫酸根、氯酸根、高氯酸根、卤离子如氯离子或羧酸的阴离子如甲酸根、乙酸根、草酸根、苯甲酸根或柠檬酸根。可以使用的过渡金属络合物化合物的实例是Mn(IV)2(μ-O)3(1,4,7-三甲基-1,4,7-三氮杂环壬烷)-二-六氟磷酸盐、[N,N'-双[(2-羟基-5-乙烯基苯基)亚甲基]-1,2-二氨基环己烷]-锰(III)-氯化物、[N,N'-双[(2-羟基-5-硝基苯基)-亚甲基]-1,2-二氨基环己烷]-锰(III)-乙酸盐、[N,N'-双[(2-羟基苯基)-亚甲基]-1,2-苯二胺]-锰(III)-乙酸盐、[N,N'-双[(2-羟基苯基)-亚甲基]-1,2-二氨基环己烷]-锰(III)-氯化物、[N,N'-双[(2-羟基苯基)-亚甲基]-1,2-二氨基乙烷]-锰(III)-氯化物、[N,N'-双[(2-羟基-5-磺酰苯基)亚甲基]-1,2-二氨基乙烷]-锰(III)-氯化物、锰-草酸盐络合物、硝基五胺-钴(III)氯化物、亚硝酸五胺-钴(III)-氯化物、六氨钴(III)-氯化物、氯代五氯胺-钴(III)-氯化物和过氧络合物[(NH3)5Co-O-O-Co(NH3)5]Cl4。
作为可以在制剂中使用的酶,除了本发明的漆酶之外,还可考虑来自淀粉酶、蛋白酶、脂肪酶、角质酶(Cutinasen)、支链淀粉酶、半纤维素酶、纤维素酶、氧化酶和过氧化物酶类的那些以及其混合物。此外,根据本发明,除了本发明的漆酶之外,还可以使用一种或多种另外的漆酶或多铜氧化酶。特别合适的是从真菌或细菌获得的酶活性成分,所述真菌或细菌如枯草芽孢杆菌(Bacillus subtilis)、地衣芽孢杆菌(Bacillus licheniformis)、迟缓芽孢杆菌(Bacillus lentus)、灰色链霉菌(Streptomyces griseus)、柔毛腐质霉(Humicola lanuginosa)、特异腐质霉(Humicola insolens)、假产碱假单胞菌(Pseudomonas pseudoalcaligenes)、洋葱假单胞菌(Pseudomonas cepacia)或灰盖鬼伞菌(Coprinus cinereus)。酶可以吸附在支持物上和/或嵌入固体材料中,以保护它们免于过早失活。其以下述量包含在本发明的洗涤剂或清洁剂中,所述量优选为至多5wt%,特别是0.2wt%~4wt%。如果本发明的制剂含有蛋白酶,则其优选的蛋白水解活性范围为约100PE/g~约10 000PE/g,特别是300PE/g~8000PE/g。如果希望在本发明的制剂中使用多种酶,则可以通过将所述两种或更多种分离的或以已知方式分开生产的酶掺入或通过将一同在颗粒中生产的两种或更多种酶而进行。
除了水之外,可用于洗涤剂(特别是液体或糊剂形式)的有机溶剂包括具有1~4个碳原子的醇,特别是甲醇、乙醇、异丙醇和叔丁醇,具有2~4个碳原子的二醇,特别是乙二醇和丙二醇,及其混合物,以及可以从所述类别化合物衍生的醚。此类水溶性溶剂以下述量存在于本发明的制剂中,所述量优选不超过30wt%,特别是6wt%~20wt%。
为了设定不会通过混合剩余组分而本身产生的所需pH,本发明的制剂可以含有体系相容和环境相容的酸,特别是柠檬酸、乙酸、酒石酸、苹果酸、乳酸、乙醇酸、琥珀酸、戊二酸和/或己二酸,以及无机酸特别是硫酸,或碱特别是铵或碱金属氢氧化物。这种pH调节剂以下述量包含在本发明的制剂中,所述量优选不超过20wt%,特别是1.2wt%~17wt%。
发灰抑制剂的任务是保持从织物纤维释放的污物悬浮在液体中。为此,大多数有机性质的水溶性胶体是合适的,例如淀粉、胶料、明胶、淀粉或纤维素的醚羧酸的盐或醚磺酸的盐或纤维素或淀粉的酸性硫酸酯的盐。另外,含有水溶性酸基的聚酰胺也适用于此。此外,可以使用除了上述淀粉衍生物以外的那些,例如醛淀粉。优选使用纤维素醚,例如羧甲基纤维素(Na盐)、甲基纤维素、羟烷基纤维素和混合醚,如甲基羟乙基纤维素、甲基羟丙基纤维素、甲基羧甲基纤维素及其混合物,相对于制剂,例如其量为0.1~5wt%。
例如,洗涤剂可以含有二氨基芪二磺酸的衍生物或其碱金属盐作为荧光增白剂,但是其优选不含用作彩色洗涤剂的荧光增白剂。例如,合适的是4,4'-双(2-苯胺基-4-吗啉代-1,3,5-三嗪基-6-氨基)芪-2,2'-二磺酸的盐或类似结构的化合物,其不带有吗啉基而带有二乙醇氨基、甲基氨基、苯胺基或2-甲氧基乙基氨基。此外,可以存在取代的二苯基苯乙烯类的增白剂,例如4,4'-双(2-磺基苯乙烯基)-联苯、4,4'-双(4-氯-3-磺基苯乙烯基)-联苯或4-(4-氯苯乙烯基)-4'-(2-磺基苯乙烯基)-联苯的碱金属盐。此外,可以使用上述荧光增白剂的混合物。
特别地,当在机器方法中使用时,有利的是将常规泡沫抑制剂加入到制剂中。例如,具有高比例的C18-C24脂肪酸的天然或合成来源的皂适用作泡沫抑制剂。合适的非表面活性剂泡沫抑制剂是例如有机聚硅氧烷及其与微细、任选硅烷化硅酸和石蜡、蜡、微晶蜡的混合物,以及其与硅烷化的硅酸或双脂肪酸亚烷基二酰胺的混合物。此外,使用不同泡沫抑制剂的混合物具有优势,例如那些由聚硅氧烷、石蜡或蜡制成的。优选地,泡沫抑制剂、特别是含有聚硅氧烷和/或石蜡的泡沫抑制剂与水溶性或分散性粒状载体物质结合。特别地,优选石蜡和双硬脂基乙烯二酰胺(Bistearylethylendiamid)的混合物。
固体制剂的生产不存在任何困难,可以以已知方式进行,例如通过喷雾干燥或造粒进行,其中任选随后加入酶和可能的其它热敏成分例如漂白剂。为了生产具有增加堆积密度(特别在650g/l~950g/l的范围内)的制剂,优选包括压制步骤的方法。
为了产生片剂形式的单相或多相、单色或多色、特别是由一层或几层(特别是两层)组成的制剂,优选地假设所有组分(任选每层)在混合器中一起混合并且通过常规压片机(例如偏心压片机或旋转压片机)压缩混合物,其压力范围为约50~100kN,优选为60~70kN。特别地,在多层片剂中,有利的是至少一层是预压缩的。这优选在5~20kN的压力下、特别是在10至15kN的压力下进行。以这种方式,随后可毫无问题地获得在使用条件下可充分快速溶解的防破损片剂,该片剂的断裂强度和弯曲强度通常为100~200N,但优选大于150N。优选地,以此方式制得的片剂的重量为10g~50g,特别是15g~40g。片剂可以采用任何空间形式并且可以是圆形、椭圆形或正方形,其中也可以是中间形式。角部和边缘有利地是磨圆的。圆形片剂的直径优选为30mm~40mm。特别地,主要通过洗衣机的配料装置引入的方形或棱柱形设计的片剂的尺寸取决于该配料装置的几何形状和体积。作为示例,优选的实施方案具有(20~30mm)×(34~40mm),特别是26×36mm或24×38mm的基面。
常规的含溶剂溶液形式的液体或糊剂制剂通常通过简单混合成分来制备,该成分可以置于所述物质中或作为溶液于自动混合器中。
本发明的固体和/或液体洗涤剂还可以例如包装在小包(优选自溶)和小袋中,特别是多腔袋中。在本发明的含义内,术语“液体”还包括在液体中的任何固体分散体。本发明的液体制剂还可以是多相的,所述相可以例如是垂直的,从而排列在彼此顶部;或是水平的,从而彼此并排。
实施例
以下实施例解释本发明,但不限于此:
实施例1:
测定具有氨基酸序列SEQ ID NO.1、SEQ ID NO.3、SEQ ID NO.5和SEQ ID NO.7的漆酶的活性。
锦葵色素()检测:通过光度测定漆酶的酶活性。为此,使用花青苷樱草素()(锦葵色素-3-半乳糖苷氯化物,CAS号:30113-37-2)作为漆酶底物。
用Synergy 2TM酶标仪(BioTek Instruments GmbH,Bad Friedrichshall,德国)测量吸收。
底物溶液由缓冲液(0.1M NaAc;0.1M磷酸钾,pH 8)和31mmol樱草素(Sigma)组成。将50μL漆酶样品加入到微量滴定板中的100μL底物溶液中并在30分钟内在30℃以mAbs/min测量吸收的减少。用水代替漆酶样品制备对比样品。
转换为单位:
使用下式计算酶活性:
活性[锦葵色素mU/mL]=(测定中的总体积μL*每分钟增加))/(测定样品体积mL*16000L/mol/cm*层厚度cm)
在微量滴定板中具有150μL容量的层厚度为0.425cm。负增加(-Abs/min)转换为正增加。
使用该方法,测定从野生型残留物纯化的漆酶的活性,其用于实施例2:
凤尾菇纯化的野生型残留物的酶样品显示出-21.4mAbs/min的活性,从而为9锦葵色素mU/mL。
佛罗里达平菇纯化的野生型残留物的酶样品显示出-20.4mAbs/min的活性,从而为9锦葵色素mU/mL。
埃杜拉香菇纯化的野生型残留物的酶样品显示出-4.5mAbs/min的活性,从而为2锦葵色素mU/mL。
栓菌属种(Tsp,DSMZ 11309)纯化的野生型残留物的酶样品显示出-29.5mAbs/min的活性,从而为13锦葵色素mU/mL。
ABTS检测:
对于ABTS检测,在MTP中加入245μL的50mM、pH 4.5的乙酸钠缓冲液,并以相应稀释度加入30μL的5mM ABTS溶液,加上10μL的2mM H2O2和15μL样品。在420nm、30℃进行10分钟的动态测量。释放的ABTS自由基的活性可以使用ABTS的摩尔消光系数以μmol/min和ml计算。
样品体积:0.015mL,测试体积:0.3mL,eABTS=0.0368cm2*μmol-1,在405~420nm
活性[μmol*min-1*ml-1]=(((DE/minSample-DE/minBlank)*p*R2)/(eABTS*测试体积))*(测试体积/样品体积)*稀释倍数
使用该方法,测定以异源方式在木霉中表达的漆酶,其用于实施例4:
来自Led的漆酶 2359ABTS U/mL
来自Ppu的漆酶 6152ABTS U/mL
来自Tsp的漆酶 3142ABTS U/mL
实施例2:
分别用以下纯化的野生型残留物之一进行第一次迷你洗涤测试:
a.凤尾菇,其中存在具有氨基酸序列SEQ ID NO.1的漆酶;
b.佛罗里达平菇,其中存在具有氨基酸序列SEQ ID NO.3的漆酶;
c.埃杜拉香菇,其中存在具有氨基酸序列SEQ ID NO.5的漆酶;
d.栓菌属种(Tsp,DSMZ 11309),其中存在具有氨基酸序列SEQ ID NO.7的漆酶。
测试条件如下:40℃,16°dH水,1小时,漆酶浓度:1锦葵色素mU/mL
对于该测试,使用标准化的受污织物。使用以下杂质:
i.醋栗CS12[CFT(Center For Testmaterials)B.V.Vlaardingen,荷兰]
ii.蓝莓CS15[CFT(Center For Testmaterials)B.V.Vlaardingen,荷兰]
iii.茶EMPA 167[EMPA: Material-und Prüfanstalt(EMPA)Testmaterialien AG[联邦材料和测试机构,测试材料],St.Gallen,瑞士]
将打孔的机织物(直径=10mm)放在微量滴定板中,将洗涤液预加温至40℃。于16°dH水中洗涤基质的终浓度为4.06g/L。
然后将液体和酶加入到杂质中,然后在40℃和600rpm温育1小时。
随后,将杂质用清水冲洗数次,放置干燥,用Minolta色彩测量装置Cm700d(CIE L*a*b*,D65/10°/SCI)测定亮度。
清洁性能以经洗涤的机织物的亮度显示。机织物越亮,清洁性能越好。测量的是L值=亮度,其越高,表示越亮。
用不含酶、荧光增白剂和染料的液体洗涤剂进行洗涤。组合物示于实施例3中。
a.用来自栓菌属种(Tsp,DSMZ 11309)的漆酶进行洗涤测试
样品1:仅洗涤剂,作为基准
样品2:洗涤剂+来自栓菌属种(Tsp,DSMZ 11309)的漆酶,1mU/mL
样品3:洗涤剂+来自平菇(Sigma)的漆酶,1mU/mL
结果(样品1比样品2)
杂质 | 样品1 | 样品2 |
醋栗 | 74.8 | 79.7 |
蓝莓 | 73.3 | 74.0 |
茶 | 73.8 | 75.1 |
可以清楚地看出,漆酶对两种含花青苷的杂质(醋栗和蓝莓)表现出良好的性能。茶渍不像用其它漆酶那样变暗,而是有利地明显变亮。性能的显著提高已可以从1个单位看出;事实上,这里已经实现了4.9个单位的改善。
市售的来自平菇的漆酶可以与样品2中的漆酶以相同活性水平作为阴性对照使用。
结果(样品1比样品3)
杂质 | 样品1 | 样品3 |
醋栗 | 76.8 | 77.7 |
蓝莓 | 76.4 | 75.7 |
茶 | 75.1 | 74.8 |
与样品1相比,来自样品3的去污力没有明显改善。
b)用来自佛罗里达平菇(Pos)的漆酶进行洗涤测试
样品1:仅洗涤剂,作为基准
样品2:洗涤剂+来自Pos的漆酶,1mU/mL
样品3:洗涤剂+来自平菇(Sigma)的漆酶,1mU/mL
结果(样品1比样品2)
杂质 | 样品1 | 样品2 |
醋栗 | 74.8 | 78.1 |
蓝莓 | 73.3 | 75.7 |
茶 | 73.8 | 74.1 |
可以清楚地看出,漆酶对两种含花青苷的杂质(醋栗和蓝莓)表现出良好的性能,且对茶而言不像其它漆酶那样变暗。性能的显著提高已经可以从1个单位看出;这里已经实现了分别为3.3和2.4个单位的改善。
对于阴性对照,参见a)。
c)用来自埃杜拉香菇的漆酶进行洗涤测试
样品1:仅洗涤剂,作为基准
样品2:洗涤剂+来自埃杜拉香菇(Led)的漆酶,1mU/mL
样品3:洗涤剂+来自平菇(Sigma)的漆酶,1mU/mL
结果(样品1比样品2)
杂质 | 样品1 | 样品2 |
醋栗 | 74.8 | 76.5 |
蓝莓 | 73.3 | 76.9 |
茶 | 73.8 | 77.0 |
可以清楚地看出,漆酶对两种含花青苷的杂质(醋栗和蓝莓)表现出良好的性能。茶渍不像用其它漆酶那样变暗,而是有利地明显变亮。性能的显著提高已经可以从1个单位看出;这里已经实现了分别为4.7和3.4个单位的改善。
对于阴性对照,参见a)。
d)用来自凤尾菇的漆酶进行洗涤测试
样品1:仅洗涤剂,作为基准
样品2:洗涤剂+来自凤尾菇(Ppu)的漆酶,1mU/mL
样品3:洗涤剂+来自平菇(Sigma)的漆酶,1mU/mL
结果(样品1比样品2)
杂质 | 样品1 | 样品2 |
醋栗 | 74.8 | 77.4 |
蓝莓 | 73.3 | 76.6 |
茶 | 73.8 | 74.3 |
可以清楚地看出,漆酶对两种含花青苷的杂质(醋栗和蓝莓)表现出良好的性能。茶渍不像用其它漆酶那样变暗,而是有利地明显变亮。性能的显著提高已经可以从1个单位看出;这里已经实现了分别为2.6和3.3个单位的改善。
对于阴性对照,参见a)。
实施例3:
洗涤剂配方:
该配方用于实施例2中。剂量为69g/17L:
不含荧光增白剂、香味剂、染料和酶。
实施例4:耐洗牢度测试仪中的洗涤测试:
测试条件如下:
i.40℃、16°dH水、1小时、200 mL洗涤液、10个钢球、16.8 g杂质和结缔组织
ii.漆酶浓度:25 ABTS U/mL
对于该测试,主要使用由果汁和果酱制成天然杂质。使用以下杂质:
i.野生黑莓果酱(Wildheidelbeer Konfitüre),Darbo
ii.巴塞尔黑樱桃果酱(Baseler Schwarzkirsche Marmelade),
iii.黑莓果汁(Heidelbeer-Saft),Rabenhorst
iv.蔓越莓果汁(Cranberry-Saft),Rabenhorst
v.蔓越橘果汁(Preiselbeer-Saft),Rabenhorst
vi.接骨木果果汁(Holunderbeer-Saft),Rabenhorst
vii.蓝莓CS15,[CFT(Center For Testmaterials)B.V.Vlaardingen,荷兰]
将杂质施加在棉无纺布(直径约5 cm)上,涂抹并储存至少1周。
在耐洗牢度测试仪于40℃运行1小时后,将杂质用水冲洗4次,然后旋转并风干。亮度在拧干后用Minolta CR400-1测色仪测量。与空白组(不含酶的洗涤剂)相比来测定颜色值Y。
对于洗涤试验,使用以异源方式在木霉中表达的以下3种漆酶:
a.来自凤尾菇(Ppu)的漆酶,其具有SEQ ID NO.1
b.来自埃杜拉香菇(Led)的漆酶,其具有SEQ ID NO.5
c.来自栓菌属种的漆酶(Tsp)(Tsp,DSMZ 11309),其具有SEQ ID NO.7
结果:
这些值表示没有酶的洗涤剂空白组的Δ-Y。认为显著的改善是只有1个单位的变化。
来自Led的漆酶 | 来自Ppu的漆酶 | 来自Tsp的漆酶 | |
野生黑莓果酱 | 1.5 | 2.2 | 2.4 |
巴塞尔黑樱桃果酱 | 1.8 | 1.5 | 2.0 |
黑莓果汁 | 0.4 | 1.7 | 0.4 |
蔓越莓果汁 | 0.6 | 1.1 | 1.0 |
蔓越橘果汁 | 0.8 | 1.5 | 0.4 |
接骨木果果汁 | 1.0 | 1.1 | 1.0 |
CS-15蓝莓 | 1.3 | 1.8 | 3.0 |
在含有花青苷的污渍上可以看到显著的清洁性能。来自凤尾菇的漆酶显示出最广泛的性能谱。
序列表
<110> 汉高股份有限及两合公司
<120> 具有改进的去污力的包含至少一种漆酶的洗涤剂
<130> PT033256PCT
<150> DE 102015210367.6, DE 102015210369.2, DE 102015210368.4, DE
<150> 102015210370.6
<151> 2015-06-05
<160> 8
<170> SIPOSequenceListing 1.0
<210> 1
<211> 533
<212> PRT
<213> Pleurotus pulmonarius
<400> 1
Met Ala Val Ala Phe Ile Ala Leu Val Ser Leu Ala Leu Ala Leu Val
1 5 10 15
Arg Val Glu Ala Ser Ile Gly Pro Arg Gly Thr Leu Asn Ile Ala Asn
20 25 30
Lys Val Ile Ser Pro Asp Gly Phe Ser Arg Ser Thr Val Leu Ala Gly
35 40 45
Gly Ser Tyr Pro Gly Pro Leu Ile Lys Gly Lys Thr Gly Asp Arg Phe
50 55 60
His Ile Asn Val Val Asn Glu Leu Ala Asp Thr Ser Met Pro Ile Asp
65 70 75 80
Thr Ser Ile His Trp His Gly Leu Phe Ala Lys Gly His Asn Trp Ala
85 90 95
Asp Gly Pro Ala Met Val Thr Gln Cys Pro Ile Val Pro Gly His Ser
100 105 110
Phe Leu Tyr Glu Phe Asp Val Pro Asp Gln Ala Gly Thr Phe Trp Tyr
115 120 125
His Ser His Leu Gly Thr Gln Tyr Cys Asp Gly Leu Arg Gly Pro Leu
130 135 140
Val Ile Tyr Ser Lys Asn Asp Pro His Lys Arg Leu Tyr Asp Val Asp
145 150 155 160
Asp Glu Ser Thr Val Leu Thr Val Gly Asp Trp Tyr His Ala Pro Ser
165 170 175
Leu Ser Leu Thr Gly Val Pro His Pro Asp Ser Thr Leu Phe Asn Gly
180 185 190
Leu Gly Arg Ser Leu Asn Gly Pro Ala Ser Pro Leu Tyr Val Met Asn
195 200 205
Val Val Lys Gly Lys Arg Tyr Arg Ile Arg Leu Ile Asn Thr Ser Cys
210 215 220
Asp Ser Asn Tyr Gln Phe Ser Ile Asp Gly His Thr Phe Thr Val Ile
225 230 235 240
Glu Ala Asp Gly Glu Asn Thr Gln Pro Leu Gln Val Asp Gln Val Gln
245 250 255
Ile Phe Ala Gly Gln Arg Tyr Ser Leu Val Leu Asn Ala Asn Gln Ala
260 265 270
Val Gly Asn Tyr Trp Ile Arg Ala Asn Pro Asn Ser Gly Asp Pro Gly
275 280 285
Phe Ala Asn Gln Met Asn Ser Ala Ile Leu Arg Tyr Lys Gly Ala His
290 295 300
Ser Ile Asp Pro Thr Thr Pro Glu Gln Asn Ala Thr Asn Pro Leu Arg
305 310 315 320
Glu Tyr Asn Leu Arg Pro Leu Val Lys Lys Pro Ala Pro Gly Lys Pro
325 330 335
Phe Pro Gly Gly Ala Asp His Asn Ile Asn Leu Asn Phe Ala Phe Asp
340 345 350
Pro Ala Thr Ala Leu Phe Thr Ala Asn Asn Phe Thr Phe Val Pro Pro
355 360 365
Thr Val Pro Val Leu Leu Gln Ile Leu Ser Gly Thr Arg Asp Ala His
370 375 380
Asp Leu Ala Pro Ala Gly Ser Ile Tyr Asp Ile Lys Leu Gly Glu Val
385 390 395 400
Val Glu Ile Thr Met Pro Ala Leu Val Phe Ala Gly Pro His Pro Leu
405 410 415
His Leu His Gly His Thr Phe Ala Val Val Arg Ser Ala Gly Ser Ser
420 425 430
Thr Tyr Asn Tyr Glu Asn Pro Val Arg Arg Asp Val Val Ser Ile Gly
435 440 445
Asp Asp Pro Thr Asp Asn Val Thr Ile Arg Phe Val Ala Asp Asn Ala
450 455 460
Gly Pro Trp Phe Leu His Cys His Ile Asp Trp His Leu Asp Leu Gly
465 470 475 480
Phe Ala Val Val Phe Ala Glu Gly Val Asn Gln Thr Ala Val Ala Asn
485 490 495
Pro Val Pro Glu Ala Trp Asn Asn Leu Cys Pro Leu Tyr Asn Ser Ser
500 505 510
Asn Pro Ser Lys Leu Leu Met Gly Thr Asn Ala Ile Gly Arg Leu Pro
515 520 525
Ala Pro Leu Lys Ala
530
<210> 2
<211> 1602
<212> DNA
<213> Pleurotus pulmonarius
<400> 2
atggcggttg cattcattgc gcttgtttca cttgccttag cactcgtacg cgtcgaggcc 60
agcatcgggc cccgcggaac gctgaacata gcgaacaaag tcatcagccc agatggattt 120
tctcgctcaa ccgtgctcgc cggtggctcc tacccgggcc cgttgatcaa ggggaagacc 180
ggtgataggt ttcatatcaa tgtcgtgaac gagctcgccg atacgtcgat gccaatcgac 240
accagtatcc actggcatgg tctcttcgcc aagggccaca attgggcaga tggccctgcc 300
atggttacgc aatgcccaat cgttccgggc cactcgtttt tgtacgaatt cgacgtccct 360
gatcaagctg gaacattttg gtatcactcc catctgggga cacagtactg tgatggactg 420
cgagggccat tggtcatcta ctcgaagaac gacccccaca agcgtttgta cgatgtcgac 480
gatgaatcca cggtgctgac cgttggtgac tggtaccacg ccccctcatt atcacttact 540
ggagtgcccc atcccgactc aacactcttc aatggcctcg gacgttccct caacggtcca 600
gcatcgccgc tatacgttat gaacgtagtc aaaggcaaac gctatcgtat tcggctcatc 660
aacacctcct gcgactccaa ctaccaattt tccattgacg gacatacctt cactgttatc 720
gaagctgatg gggagaacac acagcctctg caggtcgatc aagtgcaaat ttttgcaggc 780
cagcgctatt cgcttgtcct caacgcgaat caggcggtcg gcaactactg gatccgcgca 840
aaccccaaca gcggcgatcc cggtttcgcg aaccagatga actctgccat cctccggtac 900
aaaggcgcac acagcatcga ccccacgacc cccgagcaga acgctaccaa ccccctccgc 960
gaatacaacc tccgcccgct cgtcaagaag cccgcgccag gcaaaccatt ccccggaggc 1020
gccgaccaca acatcaacct aaacttcgct ttcgatcctg ccaccgctct gttcactgcg 1080
aacaatttca cgtttgtccc ccctactgtt ccagtattgt tgcagatctt gtcgggcaca 1140
cgcgatgcgc atgatttggc ccctgctggg tcgatttatg acattaagct aggagaagtc 1200
gttgagatca ctatgcctgc acttgtattt gctggaccac atccccttca tttacatggg 1260
cataccttcg ctgtagttcg cagtgccggc agcagcacat ataactacga gaatcccgtc 1320
cgtagagatg ttgtatccat cggtgatgac ccaacggaca acgtcaccat ccgattcgta 1380
gcagacaacg caggtccttg gttcctgcac tgccacatcg actggcatct tgacctcggc 1440
ttcgcggtcg tctttgccga aggagtaaat cagaccgcag tagccaaccc cgtacctgaa 1500
gcctggaaca acctctgccc cctatacaac agctctaacc catctaaact tctaatgggc 1560
actaatgcca ttggccgtct gcctgcgcca ctgaaggcat ga 1602
<210> 3
<211> 521
<212> PRT
<213> Pleurotus ostreatus var. florida
<400> 3
Met Val Leu Ser Thr Lys Leu Ala Ala Leu Val Ala Ser Leu Pro Phe
1 5 10 15
Val Leu Ala Ala Thr Lys Lys Leu Asp Phe His Ile Arg Asn Asp Val
20 25 30
Val Ser Pro Asp Gly Phe Glu Arg Arg Ala Ile Thr Val Asn Gly Ile
35 40 45
Phe Pro Gly Thr Pro Val Ile Leu Gln Lys Asn Asp Lys Val Gln Ile
50 55 60
Asn Thr Ile Asn Glu Leu Thr Asp Pro Gly Met Arg Arg Ser Thr Ser
65 70 75 80
Ile His Trp His Gly Leu Phe Gln His Lys Thr Ser Gly Met Asp Gly
85 90 95
Pro Ser Phe Val Asn Gln Cys Pro Ile Pro Pro Asn Ser Thr Phe Leu
100 105 110
Tyr Asp Phe Asp Thr Ala Gly Gln Thr Gly Asn Tyr Trp Tyr His Ser
115 120 125
His Leu Ser Thr Gln Tyr Cys Asp Gly Leu Arg Gly Ser Phe Ile Val
130 135 140
Tyr Asp Pro Asn Asp Pro Leu Lys His Leu Tyr Asp Val Asp Asp Glu
145 150 155 160
Ser Thr Ile Ile Thr Leu Ala Asp Trp Tyr His Asp Leu Ala Pro His
165 170 175
Ala Gln Asn Gln Phe Phe Gln Thr Gly Ser Val Pro Ile Pro Asp Thr
180 185 190
Gly Leu Ile Asn Gly Val Gly Arg Phe Lys Gly Gly Pro Leu Val Pro
195 200 205
Tyr Ala Val Ile Asn Val Glu Gln Gly Lys Arg Tyr Arg Phe Arg Leu
210 215 220
Ile Gln Ile Ser Cys Arg Pro Phe Phe Thr Phe Ser Ile Asp Asn His
225 230 235 240
Thr Phe Asp Ala Ile Glu Phe Asp Gly Ile Glu His Asp Pro Thr Pro
245 250 255
Ala Gln Asn Ile Asp Ile Tyr Ala Ala Gln Arg Ala Ser Ile Ile Val
260 265 270
His Ala Asn Gln Thr Ile Asp Asn Tyr Trp Ile Arg Ala Pro Leu Thr
275 280 285
Gly Gly Asn Pro Ala Gly Asn Pro Asn Leu Asp Ile Ser Leu Ile Arg
290 295 300
Ala Ile Leu Arg Tyr Lys Gly Ala Pro Ala Val Glu Pro Thr Thr Val
305 310 315 320
Ala Thr Thr Gly Gly His Lys Leu Asn Asp Ala Glu Met His Pro Ile
325 330 335
Ala Gln Glu Gly Pro Gly Asn Leu Gly Thr Gly Pro Pro Asp Met Ala
340 345 350
Ile Thr Leu Asn Ile Ala Gln Pro Asn Pro Pro Phe Phe Asp Ile Asn
355 360 365
Gly Ile Ser Tyr Leu Ser Pro Ser Val Pro Val Leu Leu Gln Met Leu
370 375 380
Ser Gly Ala Arg Lys Pro Gln Asp Phe Leu Pro Ser Glu Gln Val Ile
385 390 395 400
Ile Leu Pro Ala Asn Lys Leu Ile Glu Val Ser Ile Pro Gly Ala Gly
405 410 415
Ala His Pro Phe His Leu His Gly His Thr Phe Asp Ile Val Arg Thr
420 425 430
Ser Asn Ser Asp Val Val Asn Leu Val Asn Pro Pro Arg Arg Asp Val
435 440 445
Leu Pro Ile Asn Gly Gly Asn Thr Thr Phe Arg Phe Phe Ser Gly Asn
450 455 460
Ser Gly Ala Trp Phe Leu His Cys His Ile Asp Trp His Leu Glu Ala
465 470 475 480
Gly Leu Ala Val Val Phe Ala Glu Arg Pro Ala Glu Val Asn Glu Gly
485 490 495
Glu Gln Ala Gln Ile Val Thr Gln Asp Trp Arg Thr Leu Cys Pro Ala
500 505 510
Tyr Asp Gly Leu Ala Pro Glu Phe Gln
515 520
<210> 4
<211> 1566
<212> DNA
<213> Pleurotus ostreatus var. florida
<400> 4
atggtgctct ctactaagct cgctgctctt gtggcttcgc tccccttcgt tcttgctgcg 60
acgaagaaac ttgacttcca cattcgaaac gacgttgtct caccagacgg cttcgagcgc 120
agggcgatca cagtcaatgg catcttccct gggacgcccg ttatactgca gaagaacgac 180
aaggtccaga tcaatactat caatgaactg acggaccctg gcatgcgtcg cagtacctcc 240
atccattggc acggtttatt ccaacataaa acctccggca tggacgggcc ttctttcgtt 300
aaccagtgcc cgattccccc caactctact ttcctttacg acttcgatac cgcaggacag 360
actggaaact actggtacca ctcccacttg tctacgcaat attgcgacgg tcttcgtggt 420
tctttcatcg tttatgatcc caatgatcct ctgaagcacc tgtacgatgt tgatgatgag 480
agcaccatca tcaccctggc agattggtat catgaccttg ctccccacgc tcaaaaccag 540
ttcttccaaa ctggttcggt cccgattccc gacactggcc tgatcaacgg tgttggacga 600
ttcaagggcg gtcctcttgt cccttacgcc gtcatcaacg ttgagcaggg caagcgatac 660
agattccgcc tcatccagat ttcatgtcgt cccttcttca ccttctctat tgacaaccac 720
acattcgatg caattgaatt cgacggcata gagcacgacc cgacacccgc ccagaacatt 780
gatatctatg ctgctcaacg tgcatccatc atcgtccacg ccaaccagac catcgacaac 840
tactggatcc gtgcgccact cacaggggga aaccccgcag gtaaccctaa cctggacata 900
tcgttgatca gggccatcct tcgttacaag ggtgctcccg ctgtcgagcc caccacagtt 960
gcgactactg gaggccacaa gctcaatgat gccgagatgc acccaattgc tcaggaaggt 1020
cccggaaact tgggcaccgg ccccccagac atggccatca ccttgaacat cgcccagccg 1080
aaccctccat tcttcgatat caacggcatt tcgtatctct ctccttcggt tccagtgttg 1140
ctccagatgc ttagcggtgc tcgcaaacca caagacttcc tgccatcgga acaagtcatc 1200
atcttgcccg ccaacaagct cattgaagtc tctatacccg gtgccggcgc gcatcctttc 1260
catttgcacg gtcatacctt cgacattgtc cgcacttcga atagcgacgt cgtgaacctt 1320
gtcaacccac cgcgtcgcga cgtattgcct atcaacggtg gaaacacaac attccgtttc 1380
ttctctggca actccggtgc atggttcctc cattgccata ttgactggca tcttgaagct 1440
ggtctcgctg ttgtcttcgc ggaacgacct gctgaagtga acgagggcga acaggctcag 1500
atcgtcaccc aagactggag gacgctttgc ccagcttatg atggcctcgc tcccgagttc 1560
cagtaa 1566
<210> 5
<211> 515
<212> PRT
<213> Lentinula edodes
<400> 5
Met Gly Phe Gln Ser Phe Thr Ser Leu Leu Ser Leu Val Ala Leu Val
1 5 10 15
Pro Ser Ala Phe Ala Val Leu Glu Thr Thr Gly Asp Leu Val Ile Ser
20 25 30
Asn Ala Ala Val Ser Pro Asp Gly Phe Ser Arg Thr Ala Val Leu Ala
35 40 45
Gly Gly Gly Val Val Gly Glu Leu Val Thr Gly Asn Lys Gly Asp Asn
50 55 60
Phe Gln Ile Asn Val Val Asn Glu Leu Thr Asp Asp Thr Met Leu Arg
65 70 75 80
Ser Thr Thr Val His Trp His Gly Ile Phe Gln Glu Gly Thr Asn Trp
85 90 95
Ala Asp Gly Pro Ala Phe Ile Asn Gln Cys Pro Ile Ala Ala Asn Asn
100 105 110
Ser Phe Leu Tyr Asp Phe Thr Val Ser Asp Gln Ala Gly Thr Phe Trp
115 120 125
Tyr His Ser His Leu Ala Thr Gln Tyr Cys Asp Gly Leu Arg Gly Pro
130 135 140
Met Val Ile Tyr Asp Pro Asp Asp Pro Tyr Ala Asp Leu Tyr Asp Val
145 150 155 160
Asp Asp Asp Thr Thr Ile Ile Ser Leu Met Asp Trp Tyr His Ala Lys
165 170 175
Ala Glu Thr Leu Thr Phe Pro Ala Pro Asp Ala Thr Leu Ile Asn Gly
180 185 190
Leu Gly Arg Tyr Ser Gly Gly Asn Ala Thr Asp Leu Ala Val Ile Thr
195 200 205
Val Thr Ser Gly Thr Arg Tyr Arg Phe Arg Leu Ile Asn Leu Ala Cys
210 215 220
Asp Pro Asn Phe Val Phe Ser Ile Asp Asn His Thr Met Thr Val Ile
225 230 235 240
Glu Val Asp Gly Val Asn His Glu Gln Leu Val Val Asp Ser Ile Gln
245 250 255
Ile Phe Ala Ala Gln Arg Tyr Ser Phe Ile Leu Glu Ala Asn Gln Ser
260 265 270
Val Asp Asn Tyr Trp Ile Arg Ala Asp Pro Ser Thr Gly Thr Ala Gly
275 280 285
Tyr Thr Gly Gly Ile Asn Ser Ala Ile Leu Arg Tyr Ser Gly Ala Ala
290 295 300
Asp Ser Glu Pro Gly Thr Pro Ala Val Thr Ser Val Leu Ala Leu Asn
305 310 315 320
Glu Thr Gln Leu Val Pro Leu Glu Asn Pro Gly Ala Pro Gly Glu Pro
325 330 335
Glu Val Gly Gly Val Asp Tyr Ala Leu Asn Leu Ala Met Ala Phe Thr
340 345 350
Gly Thr Ala Phe Thr Ile Asn Gly Tyr Ser Phe Thr Ser Pro Thr Val
355 360 365
Pro Val Leu Leu Gln Ile Leu Ser Gly Ala Glu Thr Ala Asp Ser Leu
370 375 380
Leu Pro Thr Gly Ser Val Tyr Ser Leu Pro Thr Asn Ser Thr Ile Glu
385 390 395 400
Ile Ser Ile Pro Ser Gly Gly Leu Ala Gly Phe Pro His Pro Phe His
405 410 415
Leu His Gly His Thr Phe Asp Val Val Arg Ser Ala Gly Ser Thr Thr
420 425 430
Tyr Asn Tyr Val Asn Pro Val Arg Arg Asp Val Val Ser Thr Gly Ala
435 440 445
Ala Gly Asp Asn Val Thr Ile Arg Phe Thr Thr Asp Asn Ala Gly Pro
450 455 460
Trp Phe Leu His Cys His Ile Asp Trp His Leu Glu Ala Gly Leu Ala
465 470 475 480
Ile Val Phe Ala Glu Asp Val Ala Asp Trp Asn Thr Thr Gln Thr Pro
485 490 495
Ser Thr Ala Trp Asp Asp Leu Cys Pro Ile Tyr Asp Ala Leu Ser Ala
500 505 510
Asp Gln Leu
515
<210> 6
<211> 1548
<212> DNA
<213> Lentinula edodes
<400> 6
atgggctttc aatcttttac ctccctcctt tcactggtcg cacttgttcc gtcagctttt 60
gctgtcctgg aaacaactgg tgacctcgtc atttccaacg ctgcagtcag ccccgatggt 120
ttcagccgta ccgctgtcct tgctggcggt ggggtcgtag gtgaacttgt cacaggaaac 180
aagggcgata acttccaaat caatgttgtg aacgagttga cggacgatac catgttgcga 240
tctacaaccg tccattggca cggcatcttc caagagggaa ccaattgggc tgatggacct 300
gcattcatca atcagtgtcc catagcagca aacaactcct tcttatacga cttcaccgtt 360
tctgatcaag cagggacatt ttggtatcac agccatctcg cgacgcaata ttgcgatggt 420
cttcgaggtc ctatggtcat ctatgatcct gatgatcctt atgcggactt gtacgatgtt 480
gacgacgaca ccaccattat cagtctcatg gattggtatc acgcaaaggc tgaaactctc 540
accttcccgg ctccggatgc taccttaatc aacggtcttg gacgttactc tggaggtaat 600
gccactgatt tggctgtcat cacagtgact tcagggactc gataccgatt ccgcttaatt 660
aatttggctt gtgatcccaa cttcgtgttc agcattgata atcatacgat gactgtcatc 720
gaagtagatg gagtcaacca cgagcaatta gtcgttgatt ctatccagat tttcgctgcc 780
caacgatact cattcattct tgaagctaat caatccgtgg acaattattg gattcgcgcg 840
gacccctcaa ccggtaccgc tggatataca ggagggatca attctgccat tctgcgatac 900
agtggagccg ccgacagtga gcctggaact ccggcagtta caagtgttct ggctctaaac 960
gaaactcagc ttgtgccttt ggaaaaccct ggtgctccag gagaacctga agtaggaggc 1020
gtagattatg ctttaaatct ggctatggca tttactggaa cggctttcac tatcaatggc 1080
tatagcttta cttcaccgac ggtacccgtc ttgcttcaaa tcctcagcgg tgcggaaacg 1140
gccgattcgc ttttgcctac ggggtctgtt tactccttgc ccaccaactc gaccatcgag 1200
atttctattc cttctggagg acttgccggt ttccctcacc ccttccatct tcacggtcat 1260
accttcgatg ttgtgcgcag tgctggtagt acgacctata actacgtcaa cccggtccgt 1320
cgagatgtcg tcagcactgg agccgccggg gacaatgtaa caatccgttt cactacagat 1380
aacgcgggtc cttggttcct tcactgccac attgactggc atttggaggc cggtcttgcc 1440
attgtctttg ctgaagatgt tgccgattgg aacactaccc agactccctc cactgcttgg 1500
gatgatctgt gccccatcta cgacgctctt tctgctgacc aactttga 1548
<210> 7
<211> 520
<212> PRT
<213> Trametes species (Tsp, DSMZ 11309)
<400> 7
Met Ser Arg Phe Gln Ser Leu Leu Ala Phe Val Val Ala Ser Leu Ala
1 5 10 15
Ala Val Ala His Ala Ala Ile Gly Pro Thr Ala Asp Leu Thr Ile Ser
20 25 30
Asn Ala Glu Val Ser Pro Asp Gly Phe Ala Arg Gln Ala Val Val Val
35 40 45
Asn Asn Val Thr Pro Gly Pro Leu Val Ala Gly Asn Lys Gly Asp Arg
50 55 60
Phe Gln Leu Asn Val Ile Asp Asn Leu Thr Asn His Thr Met Leu Lys
65 70 75 80
Ser Thr Ser Ile His Trp His Gly Phe Phe Gln Lys Gly Thr Asn Trp
85 90 95
Ala Asp Gly Pro Ala Phe Val Asn Gln Cys Pro Ile Ser Ser Gly His
100 105 110
Ser Phe Leu Tyr Asp Phe Gln Val Pro Asp Gln Ala Gly Thr Phe Trp
115 120 125
Tyr His Ser His Leu Ser Thr Gln Tyr Cys Asp Gly Leu Arg Gly Pro
130 135 140
Phe Val Val Tyr Asp Pro Asn Asp Pro His Ala Ser Leu Tyr Asp Val
145 150 155 160
Asp Asn Asp Asp Thr Val Ile Thr Leu Ala Asp Trp Tyr His Thr Ala
165 170 175
Ala Lys Leu Gly Pro Ala Phe Pro Leu Gly Ala Asp Ala Thr Leu Ile
180 185 190
Asn Gly Leu Gly Arg Ser Pro Ser Thr Thr Ala Ala Asp Leu Ala Val
195 200 205
Ile Asn Val Thr Lys Gly Lys Arg Tyr Arg Phe Arg Leu Val Ser Leu
210 215 220
Ser Cys Asp Pro Asn His Thr Phe Ser Ile Asp Gly His Asp Leu Thr
225 230 235 240
Ile Ile Glu Val Asp Ser Ile Asn Ser Gln Pro Leu Val Val Asp Ser
245 250 255
Ile Gln Ile Phe Ala Ala Gln Arg Tyr Ser Phe Val Leu Asn Ala Asp
260 265 270
Gln Asp Val Gly Asn Tyr Trp Ile Arg Ala Asn Pro Asn Phe Gly Asn
275 280 285
Val Gly Phe Ala Gly Gly Ile Asn Ser Ala Ile Leu Arg Tyr Asp Gly
290 295 300
Ala Asp Pro Val Glu Pro Thr Thr Thr Gln Thr Thr Pro Thr Lys Pro
305 310 315 320
Leu Asn Glu Val Asp Leu His Pro Leu Ala Thr Met Ala Val Pro Gly
325 330 335
Ser Pro Val Ala Gly Gly Val Asp Thr Ala Ile Asn Met Ala Phe Asn
340 345 350
Phe Asn Gly Thr Asn Phe Phe Ile Asn Gly Ala Ser Phe Val Pro Pro
355 360 365
Thr Val Pro Val Leu Leu Gln Ile Ile Ser Gly Ala Gln Asn Ala Gln
370 375 380
Asp Leu Leu Pro Ser Gly Ser Val Tyr Ser Leu Pro Ala Asn Ala Asp
385 390 395 400
Ile Glu Ile Ser Phe Pro Ala Thr Ala Ala Ala Pro Gly Ala Pro His
405 410 415
Pro Phe His Leu His Gly His Ala Phe Ala Val Val Arg Ser Ala Gly
420 425 430
Ser Thr Val Tyr Asn Tyr Asp Asn Pro Ile Phe Arg Asp Val Val Ser
435 440 445
Thr Gly Thr Pro Ala Ala Gly Asp Asn Val Thr Ile Arg Phe Arg Thr
450 455 460
Asp Asn Pro Gly Pro Trp Phe Leu His Cys His Ile Asp Phe His Leu
465 470 475 480
Glu Ala Gly Phe Ala Val Val Phe Ala Glu Asp Ile Pro Asp Val Ala
485 490 495
Ser Ala Asn Pro Val Pro Gln Ala Trp Ser Asp Leu Cys Pro Ile Tyr
500 505 510
Asp Ala Leu Asp Val Asn Asp Gln
515 520
<210> 8
<211> 1563
<212> DNA
<213> Trametes species (Tsp, DSMZ 11309)
<400> 8
atgtcgaggt ttcagtctct gctcgccttc gtcgtcgcct ccctcgcggc tgtggcccat 60
gccgccattg ggcccaccgc tgacctcacc atctccaatg ccgaggtcag ccccgatggg 120
ttcgctcgtc aggctgtggt tgtcaacaat gttaccccag gacccctcgt cgcgggcaac 180
aagggtgacc gcttccaact caatgtcatc gacaacctca cgaaccacac tatgctgaag 240
agcacgagta ttcactggca tggcttcttc cagaagggga caaactgggc tgatggtccc 300
gcgtttgtga accagtgccc tatttcctct gggcactcgt tcctctacga tttccaggtt 360
cccgaccagg ccggtacctt ctggtaccac agccacttgt ccactcagta ctgtgacggc 420
ctgcggggtc ctttcgttgt gtacgatccc aatgacccgc acgcgagctt gtatgacgtc 480
gacaatgacg acaccgtgat caccctcgcg gattggtacc acactgccgc gaagctcggc 540
cccgcgttcc cactgggcgc ggatgctacc cttatcaacg ggctcgggcg ctcccccagc 600
accacggcgg cggacctcgc ggtcatcaac gtcacgaagg gtaaacgcta ccgtttccgc 660
ctggtttccc tgtcgtgtga ccccaaccac acgttcagca tcgatggtca tgacttgacg 720
atcatcgagg tggactccat caattcccaa cctctggtgg ttgactccat ccagattttc 780
gctgcgcagc ggtactcctt tgtgttgaat gccgatcagg acgtcgggaa ctactggatt 840
cgcgccaacc ccaactttgg caacgtcgga tttgcggggg gtatcaactc ggccatcctg 900
cgctatgacg gcgccgaccc ggttgagccc accacgactc agactacgcc gaccaagccc 960
ctgaacgagg tcgacttgca cccgctcgcc accatggctg tgcccggctc cccggtcgca 1020
ggtggtgttg acacggctat caacatggcc ttcaacttta acggtaccaa ctttttcatc 1080
aacggcgcga gctttgtgcc ccccaccgtg ccggtcctgc tccagatcat cagcggcgcc 1140
caaaacgccc aggatctcct cccgtctggc agcgtctact ccctcccggc gaacgcggac 1200
atcgagatct cgttccctgc taccgcggct gctccgggtg cccctcaccc cttccacttg 1260
cacggtcacg ccttcgccgt cgttcgcagc gctggcagca ccgtctacaa ctacgacaac 1320
cccatcttcc gcgacgtcgt cagcacgggg acgcctgcgg ccggtgacaa cgtcaccatc 1380
cgcttccgca ccgacaaccc cggtccgtgg ttcctccact gccacatcga cttccacctc 1440
gaggccggct tcgcggtcgt gttcgcggag gacatccccg acgtcgcgtc ggcgaaccct 1500
gtccctcagg cgtggtctga tctgtgcccc atctacgacg cgctcgatgt caacgaccag 1560
tag 1563
Claims (8)
1.一种漆酶,其包含在总长度上与SEQ ID NO.1所示氨基酸序列至少70%、渐增优选至少75%、80%、81%、82%、83%、84%、85%、86%、87%、88%、89%、90%、90.5%、91%、91.5%、92%、92.5%、93%、93.5%、94%、94.5%、95%、95.5%、96%、96.5%、97%、97.5%、98%、98.5%和99%相同的氨基酸序列。
2.一种漆酶,其特征在于所述漆酶由在总长度上与SEQ ID NO.2所示核酸序列至少70%、渐增优选至少75%、80%、81%、82%、83%、84%、85%、86%、87%、88%、89%、90%、90.5%、91%、91.5%、92%、92.5%、93%、93.5%、94%、94.5%、95%、95.5%、96%、96.5%、97%、97.5%、98%、98.5%和99%相同的核酸序列编码。
3.一种洗涤剂,其包含至少一种权利要求1或2的漆酶。
4.权利要求3的洗涤剂,其特征在于,所述至少一种漆酶的浓度相对于活性蛋白质为0.001~0.15wt%,优选为0.005~0.06wt%。
5.权利要求3或4的洗涤剂,其特征在于,其优选在5℃~95℃、优选20℃~60℃,特别优选30℃~40℃的温度范围内使用。
6.权利要求3-5任一项的洗涤剂,其特征在于,其含有额外的调节剂,所述调节剂选自2,2,6,6-四甲基-1-哌啶基氧、1-羟基苯并三唑、2,2'-连氮双-3-乙基苯并噻唑-6-磺酸酯、N-羟基-乙酰苯胺、2,5-二甲苯胺、乙醇、铜、4-甲基邻苯二酚、N-羟基邻苯二甲酰亚胺、没食子酸、单宁酸、槲皮素、丁香酸、愈创木酚、二甲氧基苄醇、苯酚、紫脲酸、酚红、溴酚蓝、纤维素、对香豆素酸、路易波士、邻甲酚、二氯靛酚、羟基苯并三唑、环己酰亚胺或香草醛。
7.权利要求1或2的漆酶用于改善洗涤剂的去污力的用途,特别是用于去除有色污渍和杂质,特别优选用于除去含有花青苷的杂质。
8.一种在含有表面活性剂的水溶液中洗涤织物的方法,其特征在于,使用含有表面活性剂的水溶液含有至少一种权利要求1或2的漆酶。
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DE102015210368.4A DE102015210368A1 (de) | 2015-06-05 | 2015-06-05 | Waschmittel mit verbesserter Waschleistung, enthaltend mindestens eine Laccase |
DE102015210369.2 | 2015-06-05 | ||
DE102015210367.6A DE102015210367A1 (de) | 2015-06-05 | 2015-06-05 | Waschmittel mit verbesserter Waschleistung, enthaltend mindestens eine Laccase |
DE102015210367.6 | 2015-06-05 | ||
DE102015210370.6A DE102015210370A1 (de) | 2015-06-05 | 2015-06-05 | Waschmittel mit verbesserter Waschleistung, enthaltend mindestens eine Laccase |
DE102015210368.4 | 2015-06-05 | ||
DE102015210370.6 | 2015-06-05 | ||
DE102015210369.2A DE102015210369A1 (de) | 2015-06-05 | 2015-06-05 | Waschmittel mit verbesserter Waschleistung, enthaltend mindestens eine Laccase |
PCT/EP2016/062440 WO2016193335A1 (de) | 2015-06-05 | 2016-06-02 | Waschmittel mit verbesserter waschleistung, enthaltend mindestens eine laccase |
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US20140356913A1 (en) * | 2013-03-29 | 2014-12-04 | Matthew Wayne Parrow | Enzymes Manufactured in Transgenic Soybean for Plant Biomass Engineering and Organopollutant Bioremediation |
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AU2016271721A1 (en) | 2018-01-25 |
WO2016193335A1 (de) | 2016-12-08 |
KR20180014817A (ko) | 2018-02-09 |
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