WO2020151157A1 - 一种降解藻类的生物制剂及其制备方法和应用 - Google Patents

一种降解藻类的生物制剂及其制备方法和应用 Download PDF

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WO2020151157A1
WO2020151157A1 PCT/CN2019/090152 CN2019090152W WO2020151157A1 WO 2020151157 A1 WO2020151157 A1 WO 2020151157A1 CN 2019090152 W CN2019090152 W CN 2019090152W WO 2020151157 A1 WO2020151157 A1 WO 2020151157A1
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protein
algae
cathepsin
preparation
biological agent
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陆祖宏
何春鹏
韩婷玉
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东南大学
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  • the invention belongs to the treatment and application field of algae (covering all seaweed and freshwater algae), and specifically relates to a biological agent for efficiently degrading algae and a preparation method and application thereof.
  • Algae such as cyanobacteria, etc.
  • cyanobacteria are primitive and ancient single-celled organisms. When the temperature conditions are good (especially in summer), they tend to multiply and grow in eutrophic lakes, reservoirs, rivers, and oceans. Drifting, gathering and other phenomena can cause serious damage to the ecological environment. For example, when cyanobacteria bloom, a layer of blue-green and fishy scum is gathered on the water surface.
  • the physical method mainly includes fishing, filtration, sedimentation, adsorption and other methods, but its scope of action is small, easy to recur, and high cost.
  • the chemical method mainly uses the oxidizing properties of strong oxidants such as chlorine dioxide (ClO 2 ), ozone (O 3 ) and potassium permanganate (KMnO 4 ) to oxidize algae in the water to achieve the purpose of rapid algae killing. The high cost is also easy to cause secondary pollution to the environment.
  • Biological law is to control algae from an ecological point of view, through ecological principles such as nutritional competition between organisms, food chain, and food web.
  • algae has high nutritional value, it has not been effectively used so far. It is mainly caused by the following two reasons: 1) because the algae contains cell walls, it cannot be directly and effectively degraded by animals; 2) the algae contains algae toxins, which cannot be effectively removed by existing industrial treatment methods.
  • the present invention provides a biological preparation for efficiently degrading algae and its preparation method and application to solve the problem of high cost, prone to secondary pollution, inability to decompose algae toxins, and inability of treatment products in the existing algae treatment process. Effective use and other issues.
  • a biological preparation for degrading algae the active ingredient is a protein specifically expressed in the epithelial cells of the amphioxus caecum.
  • the active ingredients are composed of the following proteins: cathepsin L, cathepsin B, cathepsin D, ferritin, trypsin-like serine protease, lysozymes C, lysozymes G, carboxypeptidase Z/N, tetraspanin, legumain, saposin B, subtilisin-like protease, arylsulfatase B, endo-beta-1, 4-glucanase, methionine adenosyltransferase, pancreatic lipase-like protein, plasminogen, peroxiredoxin V, VCBP4, VCBP1, VCBP3, VCBP5, Gram-negative bacteria-binding protein, alpha2-macroglobulin, chitotriosidase protein, alpha2-macroglobulin, chitotriosidase protein protein, big defensin, proprotein convertase subtilisin/k
  • the preparation method of the biological preparation for degrading algae includes the following steps: (1) Obtain the caecum tissue of Amphioxus amurensis; (2) Extract the full-length mRNAs of the caecum tissue and convert it into cDNA; (3) Use homologous recombination method or Gene synthesis method to construct a full-length functional proteome library; (4) express functional proteome products in vitro.
  • the mass ratio of the protein to the algae is 1:100-1:100,000,000.
  • the present invention uses biological agents, which greatly reduces the cost of algae degradation and treatment.
  • the biological agent used in the present invention does not produce any harmful substances and avoids secondary pollution to the environment.
  • the biological agent used in the invention can effectively remove algal toxins.
  • the invention can degrade algae into non-toxic and absorbable small molecular nutrients in a short time, and the relevant algae degradation products will be powerful low-cost and high-quality substitutes for feed ingredients such as soybean meal, meat and bone meal, and fish meal.
  • Figure 1 is an example of library preparation of co-functional proteome for algae degradation.

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Abstract

一种降解藻类的生物制剂及其制备方法和应用，有效成分为文昌鱼盲囊的上皮细胞中特异性表达的蛋白质。使用生物制剂，极大地降低了藻类降解、治理的成本。采用的生物制剂，不会产生任何有害物质，避免对环境造成二次污染。采用的生物制剂能够有效的去除藻毒素。能够在短时间内把藻类降解为无毒害、可吸收的小分子营养物质，相关藻类降解产物将是豆粕、肉骨粉和鱼粉等饲料成份的有力低成本高质量替代品。

Description

一种降解藻类的生物制剂及其制备方法和应用 技术领域
本发明属于藻类(涵盖所有海水藻和淡水藻)的治理和应用领域，具体涉及一种高效降解藻类的生物制剂及其制备方法和应用。
背景技术
藻类(如蓝藻等)是一类原始而古老的单细胞生物，在气温条件较好时(尤其在夏季)容易在富营养化的湖泊、水库、河流以及海洋中大量繁殖生长，并伴随上浮、漂移、聚集等现象，可对生态环境造成严重破坏。如蓝藻水华时，在水面聚集成一层蓝绿色而有腥臭味的浮沫等。
随着社会经济的发展，藻类水华问题已成为国内外一个主要的环境问题。藻类水华带来的危害主要表现为：阻隔空气与水中气体的交换，死亡后的藻类因为分解而消耗溶解氧，使水体中的溶解氧浓度迅速降低，造成大量水生动物缺氧窒息死亡；遮蔽阳光，不利沉水植物生长；严重影响水生生物的生存，破坏生态平衡，最终导致整个水体生态系统的崩溃；释放藻毒素；破坏水质，堵塞给水处理系统；影响景观，释放异味。因此对水体中藻类的治理成为了近几年来环保人员关注的焦点。
目前国内除藻方法大体可分为物理法、化学法和生物法三类。1)物理法主要包括打捞、过滤、沉降、吸附等方法，但是其作用范围小，容易复发，成本高。2)化学法主要是利用二氧化氯(ClO 2)、臭氧(O 3)、高锰酸钾(KMnO 4)等强氧化剂的氧化性，将水中的藻类氧化，达到快速杀藻的目的，不仅成本高还容易对环境造成二次污染。3)生物法是从生态的角度出发，通过生物间的营养竞争、食物链、食物网等生态原理来控制藻类。目前主要采用微生物防治、食藻生物、水生植物抑制等方法，这类方法虽然对去除水体中的藻类具有一定的参考价值，但并不能有效的破坏微囊藻毒素稳定的化学结构，而微囊藻毒素在食物链上具有富集的特性，因此其极可能通过食物链给人体带来潜在危害。
在饲料产业方面，藻类虽然营养价值高，但目前为止并没有得到有效利用。主要由以下两方面原因造成：1)藻类由于含有细胞壁，不能为动物直接有效降解；2)藻类含有藻毒素， 现有工业处理方法不能对其进行有效清除。
发明内容
解决的技术问题：本发明提供了一种高效降解藻类的生物制剂及其制备方法和应用，以解决现有藻类治理过程中存在成本高、易产生二次污染、无法分解藻类毒素、治理产物不能有效利用等问题。
技术方案：一种降解藻类的生物制剂，有效成分为文昌鱼盲囊的上皮细胞中特异性表达的蛋白质。
具体的，有效成分由以下蛋白质组成：cathepsin L,cathepsin B,cathepsin D,ferritin,trypsin-like serine protease,lysozymes C，lysozymes G,carboxypeptidase Z/N,tetraspanin,legumain,saposin B,subtilisin-like protease,arylsulfatase B,endo-beta-1,4-glucanase,methionine adenosyltransferase,pancreatic lipase-like protein,plasminogen,peroxiredoxin V,VCBP4,VCBP1,VCBP3,VCBP5,Gram-negative bacteria-binding protein,alpha2-macroglobulin,chitotriosidase1-like protein,big defensin,proprotein convertase subtilisin/kexin type 1，Toll-interacting protein；蛋白质的相关序列如下：
Figure PCTCN2019090152-appb-000001
Figure PCTCN2019090152-appb-000002
Figure PCTCN2019090152-appb-000003
Figure PCTCN2019090152-appb-000004
Figure PCTCN2019090152-appb-000005
Figure PCTCN2019090152-appb-000006
Figure PCTCN2019090152-appb-000007
Figure PCTCN2019090152-appb-000008
Figure PCTCN2019090152-appb-000009
降解藻类的生物制剂的制备方法，包括以下步骤：(1)获取文昌鱼的盲囊组织；(2)提取盲囊组织的全长mRNAs，并转换成cDNA；(3)利用同源重组方法或基因全合成方法构建全长功能蛋白质组文库；(4)通过体外表达功能蛋白质组产物。
文昌鱼盲囊的上皮细胞中特异性表达的蛋白质在制备降解藻类的生物制剂中的应用。
优选的，上述蛋白质与藻类的质量比为1:100-1:100,000,000。
有益效果：本发明使用生物制剂，极大地降低了藻类降解、治理的成本。本发明采用的生物制剂，不会产生任何有害物质，避免对环境造成二次污染。本发明采用的生物制剂能够有效的去除藻毒素。本发明能够在短时间内把藻类降解为无毒害、可吸收的小分子营养物质，相关藻类降解产物将是豆粕、肉骨粉和鱼粉等饲料成份的有力低成本高质量替代品。
附图说明
图1为用于藻类降解的共功能蛋白质组的文库制备实施例。
具体实施方式
以下是本发明的具体实施例并结合附图，对本发明的技术方案作进一步的描述，但本发明并不限于这些实施例。
实施例1：
1.解剖文昌鱼盲囊组织；2.提取总RNA；3.从总RNA提取mRNA；4.如图1，从3'端合成第一链，用Rnase I消化cDNA/RNA杂合体，用5'端Cap特异性抗体和磁珠法来捕获全长cDNA，洗涤全长cDNA，从5'端构建第二链cDNA，把得到的双链cDNA进行文库构建和高通量测序，利用pYES-DEST52载体和Gateway同源重组方法构建全长功能蛋白质组文库，高通量测序；5.把重组载体导入酿酒酵母(Saccharomyces cerevisiae)，海量表达功能蛋白质组，提取功能蛋白质组表达产物；6.以质量比1:1,000与太湖蓝藻混合来降解藻类，经过24小时60rpm旋转后，检测降解产物的降解状态；7.利用液质联用色谱对降解产物的降解效果进行检测，发现降解产物90％以上为分子量小于300道尔顿的寡糖、寡肽、氨基酸等小分子营养物质，未发现藻毒素；8.提取降解产物，用于饲料产业应用。相关功能蛋白质组的主要蛋白质成分为：
Figure PCTCN2019090152-appb-000010
Figure PCTCN2019090152-appb-000011
Figure PCTCN2019090152-appb-000012
Figure PCTCN2019090152-appb-000013
Figure PCTCN2019090152-appb-000014
Figure PCTCN2019090152-appb-000015
Figure PCTCN2019090152-appb-000016
Figure PCTCN2019090152-appb-000017
Figure PCTCN2019090152-appb-000018
以上所述仅是本发明的优选实施方式，应当指出，对于本技术领域的普通技术人员来说，在不脱离本发明原理的前提下还可以做出若干改进，这些改进也应视为本发明的保护范围。

Claims (5)

  1. 一种降解藻类的生物制剂，其特征在于有效成分为文昌鱼盲囊的上皮细胞中特异性表达的蛋白质。
  2. 一种降解藻类的生物制剂，其特征在于有效成分由以下蛋白质组成：cathepsin L,cathepsin B,cathepsin D,ferritin,trypsin-like serine protease,lysozymes C，lysozymes G,carboxypeptidase Z/N,tetraspanin,legumain,saposin B,subtilisin-like protease,arylsulfatase B,endo-beta-1,4-glucanase,methionine adenosyltransferase,pancreatic lipase-like protein,plasminogen,peroxiredoxin V,VCBP4,VCBP1,VCBP3,VCBP5,Gram-negative bacteria-binding protein,alpha2-macroglobulin,chitotriosidase 1-like protein,big defensin,proprotein convertase subtilisin/kexin type 1，Toll-interacting protein；蛋白质的相关序列如下：
    >cathepsin L
    MRLLIAVVCVAMATASPLMNPQWEVFKKAYNRVYAAEEEFARRLIFEDNLKTIQMHNEEADRGLHTFRLGVNQYADMTHKEFLENVIGGCLLDTNTSKSTADHVHEYDPTLTDLPDTVDWRDKGYVTPVKNQEQCGSCWAFSTTGSLEGQHFKSTQKLVSLSEQNLVDCSRKRGTGLPGRLMDQGFKYIKDNGGIDTEECYPYKAKNEKCNYQASCSGATLTAKRRQDEGRGALQQAVATVGPISVAIDAGHSSFQLYQSGVYHKFFCSETKMDHGVLAVGYGTEEGKDYWLVKNSWGASWGEKGYIKMSRNRHNNWGIATSASYPTV
    >cathepsin B
    MLAWVVLSVLAAVSAKEFPIHQPLTQEIIDYVNTIDTTWKAGWNFQGATVSYVKGLCGVIRDPNNHKLPLKLHELNAQDIPDTFDSRTQWANCPTIKEVRDQGSCGSCWALAAVEAMSDRICVASKGSTMAHISAEDLNSCCKSCGNGCNGGFPEAAWEYWKRDGLVTGGPYGSHQGCQPYEIKPCEHHINGSRPACGKLEPTPRCKKSCESGYNVTFAKDKHYAKTAYSVSSKVQQIQMEIMTNGPVEAAFTVYADFPHYKSGVYQHESGAELGGHAVKMIGWGTEGSTPYWLIANSWNTDWGNMGFFKILRGQDECGIERDIVAGEPKLD
    >cathepsin D
    MKFLSVLFALVVFASALHRIPLTKMKTVRRQLADVGITYDQVLDKDYSGKYYNIKDAPEPLTNYLDAQYYGPISIGTPAQNFQVVFDTGSSNLWVPSKKCKLSDIACLLHNKYDSTQSSTYMKNGTDFAIRYGSGSLTGFLSEDTVTIGGLKVQNQTFAEAVTQPGITFVAAKFDGILGMGYDTISVDGVVPPFYNMVQQKLVDKPVFSFYLNRDPSSTTRGELLLGGTDPKYYTGDFTFLDVTKP GYWQFKMDGIMINGKATDYCKGGCAAIADTGTSLIAGPTTEVQALNKQIGATPIPGGEYMVDCSQVSSLPPISFMLGGKAFELQGKDYVLQVTTMGQTVCVSGFLGIDVPAGPLWILGDVFIGPYYTLFDMGNNRVGFAPTAHVNTTRV
    >ferritin
    MHAMMKTLTFVLLVLQCTLGARREDNQWLISREDAGDPDNQDFARQNFHEDCEAGINKQINLEMFAGYTYRSMASYFNRDDVALKGVADFFRHHSEEELEHAQLLEEFQNKRGGRVVYENLRKPEKDTWGSALEAMQAALTLEKNVNQRLINLHKTAGQHNDMQMQDFLDSHFMTEQVEGIKQIADYITNIKRVGTSGWASTCSTSTPSAAASRGNSVRRTREERNLHRTKSYL
    >trypsin-like serine protease
    MIWCLLLLALAARGDAWLFSCSASDFQCSNGNCIPGSWECDGEADCSDGSDEASCSGGGTCQQGVHFFCANGLYCIDSNWVCDGDNDCGDMSDEQNCGGGSTGTTGTTAPITSFTGDCGQPAISPQNVRVVGGVQAVQGSWPWQASLKLYGGHVCGGQIIAPNWIVTAAHCVDGQSNPSQWRVSLGSHRRTSTDSTQQDFSVTRIIMHESYDSNRINNDVALMKLSGNAQFNNYVSPICLPTQDVAAGTNCVTTGWGDTGSGASTYLMQATVPIMEWNKCNSAQYMNGAITDKMICAGYDQGGKDACQGDSGGPLVCNYSGKWTLDGIVSWGYGCAQAYKPGIYTRVTQFVSWINNKMASN
    >lysozyme C
    MFLAVVLVMGLVCSAHAKTYERCELARELVSRGLTSRSQAGEWICLVQHESSFRTGARGGPNWDGSYDHGLFQINDHYWCDNGGPHNDCGVSCSNLRDNNIADDVRCAKLIYQRHGFSAWYGWQNNCQGSTSSYVSGCF
    >lysozyme G
    MMFLVILAIVGVATADDWQCTTSVAANGQNGQCAHVDTCPYHYFVSNKCPSYGNDVKCCYNCHLGGCSISGGSSGGTGGSGSGSYGNIMEVDTTGASSQTASQDNIWYSGVSASHQLASNDLGRLNNYKSQIFDAGNAKNMDAAVIAAIISRESRAGAALAADGTGDNGNGFGLMQVDYRYHTPAGGPYSTEHMMQGTQILIDTINCVKRNHPNWNDNMTLKGGISGYNAGCGNVQTYAGMDGGTTGEDYANDVVARAQWLKGQGF
    >carboxypeptidase Z/N
    MIFAILLAALPALALAEVDSSFVFEYHRYTALRSVLLAVSQDCPDITRLYSIGQSVQGRELLVLEISDNPGQHELGEPEFKYVGNMHGNEVRGRELIILLAQYLCGEYKAGNSRIVSLVRDTRIHLMPTMNPDGFEVAANQGPDNNGWTTGRNNMQGIDLNRNFPELNSIAYSGESSGTNQDHIPIPSSYWSGTVAPETRAMITWLQSYPFVLSANMHDGDLVANYPYDTAKSGGFWGSGYAATPD DALWRDLASTYAQAHGTMATTGGGSCGFQGQGGITNGADWYSLSGGMQDFNYLHTNCYELTLELGCDKYPRESELRMEWNNNKESLLAFMEKVHIGIKGVVTDTNGNGVADAKIKVQGNAHGVNSAADGDYWRLLRPGTYSVTATKGQASQTKSCTVGSGAATTCDFTLDAAGVLSGGTTGTGRLDISM
    >tetraspanin
    MEAKGGMQCIKYLLFAFNLIFWITGLAMIGVGATVKIAFGDYFDFTGNAFASAPVFLIILGVFVSIIGFLGCCGAIKENYCMVTTFAGRLSLIFILEIAAGITGYVLRNDIENVIDTAMTDFMMKWTNKTEGRLAFDGVHKKFECCGVTNYTDWFDKSPNFNPNNQPNVPGSCCLNAKTEFDVKNPDDTCGEGADTTKINTEGCLLKLEGWLAQNILIIGGVGIGIAFIQVVGIIFACCLMRSIKQEYEVV
    >legumain
    MLSIVMLSLFGAVLGFPADFEEAIPVAEPEPGVNWAVLIAGSTGWGNYRHQADVCHAYQILHRNGIPDERIVVMMADDLAHNIRNPTKGIIINHPDGKDVYHGVPKDYTRFDVTAKNFLRVLKGDKEGVAGIGSGKVIESGPHDNVFVYYTDHGAPGIVAMPHGGMLHADDLVTTLKEMHQENKFNKLVFYLESCESGSMFDKMLPENINIFATTAANGKESSYACYMDTKRKTYLGDLYSVNWMEDSDVEKVDKETLEQQFEKVKMLTNRSHVQEYGDVNMKNDVIGLYQSTINYPSTPLPRYKQVPHDAVPAPDVPLAILNLRLETAETEEERQTVLAELKEELNLRTTIAQTMKDIATMATDDADKAEDILVNHHRDITARACYKAAVNYFHDKCYDLSQYEHALRHMYTLVNLCEEQVPLDRVTAAMDSVC
    >saposin B
    MKSVAVVLAVLLASQCLAVRPPSRRGPALVRAGGLSIGQVFKPSKVDLDLCKFCIDFSQQALNQLLNIILQVGVVGTCGDICQALADKTGSQALGVACNLLCDIEGVKEFVKIIENADLDPIYYCELLKQCTIKDDGDAKITSLDVSPKSGPQGTFMIDIEYMSKNGTGTGEMVVDIDTVDGIPLGQGFILEAQEAGSYGQKISLKAQPDPDCDPTQGPCEQWMPGVYNVNIAICNGECGSKHPHSQIYDQGMTNFTITG
    >subtilisin-like protease
    MKSLRVLAVLAAVLGVAMATAPMLRLDSKTAIPNEYIVVLQKNLTNYAVGKHMNSVKTLLTGTNDTKILFEHNFRWFKAYSIRTNQKMIRHLAEQPEVRYIEANQVVKAYQAQCQDQLEATWGLVRTVERDLLLDGIYHYQGGDGEGVDAYIIDTGIYTEHSEFGGRAKWGVDTVDTPSPETDENGHGTHVAGTVMSDAWGLAKKATAIAVKVLSRSGSGSTAGVIEGVEWTGQQHSGNNKKSVANMSLGGGRSDAMNEAVKAVVEAGVVMVVAAGNEGWEACNVSPASEPTAITVGCSD NEDDFCFFSNYGTCMDIIAPGQDVTSAWIGGQFADNTISGTSMSAPHIAGIVAKYMSRQSSVPSPEEVKDFLQTTSTKDKINQIPSNSDTLNYLGFMDCGGGPDV
    >arylsulfatase B
    MLRVPLLLLTGALVGLVSAQDGSQNATKPHILFIVADDLGWNDVGWHNPDVKTPVLDKLAHEGVILNQSYVNYVCTPSRTAFMTGYYPYHAGSQHLVFLPQQAQGIPYNFTFLPEKLKDLGYATHMVGKWHLGFCNWKYTPTYRGFDSFYGYYNADEDYYTHVVAGGLDLRDDKEVVNTKNGQYGTYFFTDRMVDIIEKHPADTPLFLYLPFQNVHEPLEVPERFENIYMNVQNENRRKLLGMVSALDEAVGNVTMAMKNAGLWDNTLVIFTTDNGGWIIASGNNYPLRGGKVTLWEGGTRGVAFVHGKMLQKTGYTNNEMIHAVDWFPTILAAAGGTADDAMDGVSQLDTIMSGQPSPRTEFVYNIDDVFPTKQGAIRIICTLCLRVGDYKLIEGVAGKPDGWIHPDNLTDSQLDSGDPTSGTWLFNLKDDPTEHHNLADSMPDKLKEMQAKLAEYRKSLVPAINPPPDPKSFPKNWGGAWSPGWC
    >endo-beta-1, 4-glucanase
    MDQDYERRLLRQQNGQNCFGSPVSSPISPMAGRESLGTSPVSSPSKDKYGDRFIPSRAGANWEIGFNSIQGMYEKTSGQARKAREANSDNGKDGLAYTCLLKNELLGAGIEDLKEQTEDRRGVLSPTTPEKRNLFRYHLTAKQASPENTDHLSPYSLSPVGKKSQKLLRSPRKQTRKISKIPFKVLDAPELQDDFYLNLVDWSATNILSVGLGTCVYLWSACTSQVTRLCDLSCDGDSVTSVNWNERGNLVAVGTHKGYVQVWDAMAGKRISMLEGHSARVGALAWNADILSSGSRDRLILQRDVRTPSVVPERRLAGHRQEVCGLKWSPDHQHLASGGNDNKRIVYKKGTFVDSFRPTNMRCTALILLALLGAARAQSYYEVTGDWGDRFQANVYIPVSQAVNGWTATLKFDKAVELEIWLAEVTSTSDGNTVFELKDMGWNANVPAGTFELSFIANTQGSAANLVGLWFNGQEVTSGGTGTGTGTGTGTGTTHRPSTVSTSHTHATAPTAAQSGTTLGVVPPVVGAYDYVDALAKSILFYEAQRAGNLPGNNRVPWRRSCCQGDGQDVGLDLSGGWFDAGDHLKLHFPLSYTITVLSWGMIEFKRAYEAAGEMANALDSIKWVTDYLIKCHPSKFEFVAQVSTHGYSQNQILVWKYPSLVQVAKLTGHSYRVLYLAMSPDGEAIVTGAGDETLRFWNVFSKTRSNKESKSVLNLYTHIR
    >methionine adenosyltransferase
    MANQDTFLFTSESVGEGHPDKLCDQISDAILDAHLKKDPNAKVACEACTKTGYIMVFGEVNTTATIDYQTVVRDVIKRVGYDHSDKGFDYNTCAVIQAVEQQSPDIAQGVHEKRGAEDIGAGDQGLMFGYASDETEECMPLTCMLAHGLNRKLAELRRDGTFTWLRPDTKTQVTVEYRMSGGACVPIRVHTVVMSVQTDDVVTLDDLREQLKERVVKAVIPSRYLDDKTIYHLQPSGKFQRGGPQADAGLTGRKIIVDTYGGWGAHGGGAFSGKDFSKVDRSAGYAARWVAKSLVKAGLC RRVLVQVSYAIGISHPLSVLVNSYGTCQDGMTDARLLKIVNKNFDLRPGVIVRELGLQRPIYEKTACYGHFGRMDPDFTWEQPKKLDLNV
    >pancreatic lipase-like protein
    MLLSVLSILSLACAAQAAEVCYGDLGCFTNIAPFWGLNRPLSSLPDPPDVVGTKFYLYTRANPTMALRERLVADSIATLSASHFSSSKPTRMVIHGFTGSAEHAWVQTIVDELLLKEDINVITVDWADGASIGGSYGQATANSRVVGAEVAKIVNYMSAQTGANARNFHLIGHSLGCHVAGYAGDILGNVGRITALDASEPYFDGMDAIVKLDPTDALFVDVIHSDGSPFIGTLGMGTSLPTGHVDFYPNGGEYQPGCHDNFVSSVVSTGFGLLTEGYDGAEAAAACSHLRAIDYFTESINSECPFTAYPCESYDKFKDGFCMSCAGSTCSQMGYRARDHYGARGKLYLTTTDGTQPGGKFCAYHYKVQVTSSTRMEETAGRIYVTFQGPSGITDTMEVTDGDVMDLKPGVTYQKLVSSASNIGDVSRVGLRFERSSSIWDFFAAQYYTVGKVTITAGETQTSYKFCGSDTPVNNGETLQLTQVGGTC>plasminogen
    MRTLLALLGLLALTAAQDCYVGRGSAYRGAVNVTSDGLECQPWSVQAPHGHAFTPANYPDAGLDGNYCRNPDNDLRPWCYTMDPNVRFGYCDAPQCDCLFNTGEFYKGSKATSISGLTCQRWDSQSPHQHDRTPQNYPEHDLRENYCRNPDGEPRTWCYTTDPNVRWEHCDVPKCDTADLGTCGVPAIQPTFSRIVGGSVATPGSWPWQVSLGYGSNGQHVCGATLIAPEWVLSAAHCFAQLSTNPGSFIVKVGKHNKASTDSTEQRMQVAQIIVHPRYQQDGQNTNDIALLKLTQRVQLNDYVSQACITETEAPEGAICVATGFGNTEGTGGDNYLKQVQLPSLSNAQCRSWLGSVIKSTMVCAGYEGGGSDTCQGDSGGPLTCPRLGQWFVSGVTSFGQGCADPRKPGVYTRVGYYIDWILKYTADN
    >peroxiredoxin V
    MLIPVTICSLARRLPSAIGARCAIYTATAYNMPIKVGDKLPGIDLYENTPGNKVNVSELFAGKKGVLFAVPGAFTPGCSKTHLPGYVGKAGDLKAKGVQVIACVSVNDPFVMEAWGKDQKAEGKVRMLADTGAEFTKAIGLDLDATGLLGNIRSKRYSMLVEDGEVKQLNVEPDGTGLTCSLAEGLKL
    >VCBP4
    MAPFTPFVLCIILAGAMAQSPFRIMTVTVPRPEVTANVGSDVKLPCFYSIPAASSGLNPTINWYKGDEGIASATKIFSLSYHGSTQIAEAYEGYELRVELESLTDPTLKLSGVRASDHGRYWCAVFDSDEHNQFGMDSKSVLLTVIDPSIQQDQGMGRPGRVELRVPAVRQVYQGEDVELTCECDDCHWTSTKSWFTVSFDDTWAATEELVATVGPCVSHQCHGGTTTVAPAFRDHFSATREALRISAAKRLDKRRYWCQVEDMTFSRGSGVDAQSVALAVESRCEGKPAGRYQHPDDCSKYYTCA EGGLQYDGISACPPGLMYDQANGYCNWATQVTCL
    >VCBP1
    MKFVLGLVLLAVGAHAMTIVTVSTPEPKVEASVGGSAELKCEFDIQPNSTQPPTIAWFKGNDDFRGVERIYTGHKVWGNETERREDSFGDYIGRVEVADLDKPAIKISGIKSTDFARYWCTVAEWGVRTELGVDAKSVLLTETGHSEASIDISVSGEKDVEEGGDVEMTCRCHGCTSAAIFDWFKGAFAGSEWVTTGNYTHIAAKVDVGVLGFPNPIEIDDGFGQFSVTPSNSLRLTGAQVADAGRYWCKVTSGGSVDIKATVLKVKVPEFTCAGKADGYYPDPEDCAMYYQCLYGFPQPFHRPCGYAGMVFNPEHLYCDWAFNVGPPCGSKA
    >VCBP3
    MQMFLLVSVCLGMAYGQSIMTVRTTHTEVEVHAGGTVELPCSYQLANDTQPPVISWLKGASPDRSTKVFKGNYNWQGEGLGFVESDSYKESFGDFLGRASVANLAAPTLRLTHVHPQDGGRYWCQVAQWSIRTEFGLDAKSVVLKVTGHTPSNNVHVSTAEVVQVDEGNDITMTCPCTDCANANVTWYTGPTFFENYETGTYQPLANKNQFGITWFSSEIAGRASFSGARNLVIRAAKITDAGRVWCELATGQGELDADRSSTILKVQLEPFTCDGKPTGLYADPTACDYYYQCIPGYPPLHRPCGYAGMVFNEEMQYCDWDINVPPPCGSKPV
    >VCBP5
    MSVLLVSIILLLYVGPESADAVSITTVTVPDRGGWVVMWPRPGDPTWVNRIEFRCEYSISPASATPPTITWLKGVFADADRQVIYKWSSSGEVYVHPEYAGRVSVPSRTHPTLVLTDAKFDDWGRYWCRVTNEDQSDEFGTDEESLLFWYKLGYDPPTRLSQVSLDKTPVRVATGGTARLDCTGTSGREASILWVKGPTSCTQGESCDSYETVIHKSAVWGAGNPEPENITISPSFDGRVSLDADGSGSFTPTLTITDIRPSDSGRYWCAPDISEVYSNLGLLNRDAQSVVIIVNDPGTEPTCAGKPDGMYQHPADCAQFYTCSGGLSYGTNNCPAGLVFNQELQLCDWANNVICL
    >Gram-negative bacteria-binding protein
    MALLAVLLFHAALWAAGEAQGTYLGCYIDTPNDRDFPHVTTMTYTNGYTTCSQYCASLGYSYIGLQYAIECFCGHTYGKHGRAAESDCYYGCSGGTGGKCGGDYRNTVWDIGDVGGGSTGTGTSDNLELVFEDTFDTLDTTKWRPEVTAGGGGNEEFQYYMGHPDNLYVQNGNLYLKPTLASEHYGDNFMTSGYIDFWHGYNYVGCFNDGGDADLTESLLTASDMNPGKCVTHCANGGYEYFGLKNTECRCGNSFGRFGHGHGCHISCPGDSGKECGGNGVTSVYGQTYGDCTSDFPQGRACYLQGSQNAPLPPIQSARVTTVESFAFKYGKVEVWAKLPTGDWIWPAIWLLPRDNVYGGWPASGEIDIMESRGNRNLYGSWGGSIGVDDMGSTLHWGPRWPYNGFQQTTASKVEVDDQEVLLVSPPTNFWDMGNFGLPSYENPWAGGGKMTPFDREFYLIFNVAVGGTNYYFPDD ATNQPYPKPWSNNNSREGAMRDFWNAKNQWYPTWNGDDVAMQIQSIKVWRFV
    >alpha2-macroglobulin
    MRGLLLLCGVLGSVPAQKLDPTPGPTPGTPGTFLIMAPKVVRPGRPLTLSVSILDPAAASVQVTATLLQNGRPTGTPVSGDFSRNADGTLDLVVPGALTAGSAYKLKVEGTGGLTFTEEKDLTFNSKSSSIFVQTDKGKYKPGQTVKMRALAVDAELKTKKEPFDIDIFDPQGNEIVQWRGLQNDDGLVEKEFALSDQPLLGDWKIVAMLPGGDEVEQPFGVEEYVLPKFEVTISTNSFFARTEDTLQGTFNAKYTYGKNVEGQGKVTVKLRSVGSNFRPTDNSIEHNNIAMVEGSGSFQFTRGDIEGLYGNPDQLYYGSYQLELIVEVTEGLTGNTQEATKELDLVQTKYKLESFNTPDNFKPGLQYTAYVRTSYHDGTPMLGMDLAQPVTLSKNYGWDGPTAPPAEELTIPGSGVIKIVMDAPSTANSMTLSVMYNDISETATTHTAQKAESPSSSYIQVTTSTPEVTAGEDAQFTVKSTSPIGYFSYQVLSRGNIVVVDRVEVTPEDTLKSFTVPTNTQMAPSSRMVVYWMTDTGEICNDGLTFTVKGAFENQVSLSYDKDRAEPAEDIAVTVRADPDSLVALLTVDQGVLLLQSGNDITQDKVLAELEEYDPAEDTSNSGGPEIMWRRRKRSIWWPMPTSGDDAHSVFENAGVIVLTDAMVYQKQPDYWWQWGILPVADFDLGMMEFMPAAAAGESAGGGGGRGGLQEVKRVRSFFPETWLWTNATAGADGTAVFASTVPDTITSWVASAFALNDVSGLGVADTPASMEAFRPFFVSLNLPYSVVNKEETAIQALVFNYMDVDMDVTVTLKGSSGFMHIVMDTNDVNSIGTEVAGDQAVVLTVPAGSSRSKSFPIIPKQFGAITVKVLAQSTLAADAVERQLLVEPEGEQKEYTKSFILDLGDSGNGDITKDLYLSVPQSGLVAGSAYARVSVMGDMMGSTISGLDSLLQMPYGCGEQTMITFAPNVYVMHYLNETNQVTPEIKSKALKFMTSGYQRELTYTHDDGSFSAFGSSDASGSTWLSAFVMKSFSQAMDYIYIDPKVLAKTIRWLIRQQQDTGVFSEPGNVIHKEMQGGLSSDVTMTSYVLVALMEYDKILATLDAPDSDKVDVTIAKNRAVAFLESRLDSETDVYTIVTMTYALTLAGSRRKDAAFQKMNSLATTEAGQMYWTKGESATSPPVSDDLMWTPPYHNPPSADVEMTAYALLTYLDRDDLVAGMPIMKWLTEQRNAYGGYSSTQDTVVGLQALGYFAAAVRAGGLDMTVTITSDTDTDFSHTFTVNDQNAIVLQRVEIPNVSGTLTARAQGQGMGLLQLFVRYNVDVVEAVPAFNLIATVKNSDTNSVTCETCGTYQLGGSSGMAVMEIGIPSGFYVEDADLMALVDNQAMPTLKRAERSTDNKRAVIYFDEMNASPTCVTLTSHRSMPVAKTQPSVVKVYDYYKPDVKASNIYQSETLKNTNICEVCADCTGCSGRQGRPRSAAAGLAVDTLLGVVLLLLTRLLVRQCRPTNTCEECLNVPCSELVTKLRL
    >chitotriosidase 1-like protein
    MLTVLALLCVMLSLGQASVIPAQRNTYRRVCYHTNWSQYRHGVGQFFPEDIDPTLCTHAIYAFAKMTNNQLQPYEWNDDSTDWSTGMYERFNTHLKPHGVKTLLGVGGWNFGSTAFSDM AATAAGRRTFVTTSIDFLRQRNFDGLDLDWEYPAARGGRPSDKQTFTLLRKELREAFDAEGARTGRAPLLLTAAMPAGEDNAMNGYEMAEVAGYLDFLNLMAYDLHGQWETYTGLNSPLYAAGDETGDDRKLNQAFAVDMWLAGGVPPSKLNLGMGTYGRSFTTTGDNSIRAPANGGGNAGLYTREKGYLAYYEICTMLNQGATRVFHSEHLSPYAYLGNQWVGYDDVESLTYKVEYLKSKNLGGAMVWAYDIDDFSGSSCGQGRYPLMNAIKNLLETGSAGVVLPPGPTHPPINPATQAPNPGVTQGPNPVVTQAPVVVEQGNLAAVTCDSKPDGFHPDPTDCGKYFQCWGGTMWPGHCSNGLQWNQAMLGCDWPYNVNC
    >big defensin
    MEKKTAYCLLFLVLLVPYTALGAVLKRAPAKKEKRAVPLAVPLVYWGASVSPAVWNWLLVTFGAAAVAAAAVTVSDNDSHSCANNRGWCRSRCFSHEYIDSWHSDVCGSYDCCRPRY
    >proprotein convertase subtilisin/kexin type 1
    MGRFYAWLVLAIVAFSSHGRAEDGEDQPGHFTNTWAVEIYGGPDRADLLALDHGYENLGQIGNLEDHYLFRHKDVPHRSRRGAHQHTKRLGDDERIQWVAQQVGRARSKRGPMGQQRRQSDDTRPMTFRDPYWEKQWYLHDTRTSTNLPKLDLHVLPVWRKGITGKGIVVAVLDDGIEKDHPDLVDNYDPDASYDFNDNDDDPQPRYEETNENKHGTRCAGEIAMAANNSECGVGIAFNARIGGVRMLDGVVTDAVEASSIGFNIQHVDIYSASWGPNDDGKTVEGPEKLARAAFEKGVREGRGGKGSIFAWASGNGGSNGDNCDCDGYTSSIYTVSISSASQQGGSPWYGEKCASTLATAYSSGEYKDQKISSTDLHHECTDSHTGTSAAAPLAAGVLALALEANPNLTWRDVQHLIVWTSEYDPLSSNPGWFQNGAGLWVNSRFGYGLLNAEAMVDMALTWKTVPEKTKCEVRIENFQPRDLGNGEEIIIELETDGCRGQNHVEALEHVQVKTTIDYSRRGDLRIVLTSPSGTSTTLLDTRRQDKSQMGFQDWPFMSTHNWGEKPQGKWTLTIEDKSDHAENNGVVKDVVLILHGTPEQPAYQSGGRVYTDYNRVQGDRNVEISAAKAAPAPAADAAPARGSEERIPGSGPFGSAASVVIEEIPEKETEFLVNWQDGMNRYETDFNPVNSGPFEPSADAGSDVYMDAEDLRPWEAFKYLQRQMKSPSGQRHPAHAYNKPSSQYIRDPWVSQNALDKEANLVKYYLQLLGYES
    >Toll-interacting protein
    MATKVTKRGQVMLGDLPDDFLRIEATPQQLQATADMQAAQMYAQYGFMGQSSGRLSITVAQAKLAKNYGVTRMDPYCRIRVGHAVFETPTAYNGAKNPRWNKVVHAPIPQGVNSFYLEIFDERSFTMDDRVAWGHITIPESVFNGETVDDWYTLSGRQGDDKEGMINLVLSHSTVPMMQQVPQVMPQMPMMMVPQPVYYPTVQAGVPYQVPAGQMPAQQQPPAQPAITEEDLKALQDMFPNMDAEVVRSVLEANRGNKDAAVNSLLSITAE
  3. 权利要求1所述降解藻类的生物制剂的制备方法，其特征在于包括以下步骤：(1)获 取文昌鱼的盲囊组织；(2)提取盲囊组织的全长mRNAs，并转换成cDNA；(3)利用同源重组方法或基因全合成方法构建全长功能蛋白质组文库；(4)通过体外表达功能蛋白质组产物。
  4. 文昌鱼盲囊的上皮细胞中特异性表达的蛋白质在制备降解藻类的生物制剂中的应用。
  5. 根据权利要求4所述的应用，其特征在于所述蛋白质与藻类的质量比为1:100-1:100,000,000。
PCT/CN2019/090152 2019-01-24 2019-06-05 一种降解藻类的生物制剂及其制备方法和应用 WO2020151157A1 (zh)

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