裂解酶以及不对称合成(S)‑苯基乙酰基甲醇的方法与流程

(S)-苯基乙酰基甲醇是有机合成中有价值的手性单元并且可用于合成精细化学品和药物。根据迄今的现有技术，并不已知能够借助不对称合成由非手性的廉价化合物以>89%ee的光学纯度生成(S)-苯基乙酰基甲醇(S)-PAC的方法。但是，在制备精细化学品或药物时，高的光学纯度是极其重要的。根据现有技术已知制备(S)-苯基乙酰基甲醇的各种方法。一方面，已知化学合成。基于化学不对称合成的(S)-PAC制备方法生成68%或86%的ee。这些方法描述在出版物Davis,FranklinA.;Sheppard,AureliaC.;Lal,G.SankarTetrahedronLetters,1989,第30卷,7第779-782页和Adam,Waldemar;Fell,RainerT.;Stegmann,VeitR.;Saha-Moeller,ChantuR.JournaloftheAmericanChemicalSociety,1998,第120卷,4第708-714页中。此外，还有仅作为副产物产生(S)-PAC且(R)-PAC以对映体过量形式存在的方法，这例如通过下列反应，例如描述在出版物Toukoniitty,Esa;Maeki-Arvela,Paeivi;Kuzma,Marek;Villela,Alexandre;KalantarNeyestanaki,Ahmad;Salmi,Tapio;Sjoeholm,Rainer;Leino,Reko;Laine,Ensio;Murzin,DmitryYu,JournalofCatalysis,2001,第204卷,2第281-291页中的1-苯基丙-1,2-二酮的还原和描述在Fleming,StevenA.;Carroll,SeanM.;Hirschi,Jennifer;Liu,Renmao;Pace,J.Lee;Redd,J.TyTetrahedronLetters,2004,第45卷,17第3341-3343页中的由苯甲醛起始的合成和描述在Brussee,J.;Roos,E.C.;Gen,A.VanDerTetrahedronLetters,1988,第29卷,35第4485-4488页中的2-羟基-2-苯基乙腈的反应。此外，描述了手性单元1-苯基丙-1,2-二醇可以氧化成(S)-PAC的合成。(S)-PAC以91%的对映体过量产生，如由Zi-QiangRong,Hui-JiePan,Hai-LongYan和YuZhaoOrganicLetters,2014,16(1),第208–211页描述，或69%的对映体过量，如由WaldemarAdam,ChantuR.Saha-Moeller和Cong-GuiZhaoJournalofOrganicChemistry,64(20),7492-7497;1999描述，但是此外被必须复杂分离出的区域异构体污染。此外，已知描述在ÁlvaroGómezBaraibar于2013年的题为“Developmentofabiocatalyticproductionprocessfor(S)-alpha-hydroxyketones”博士论文中的酶促不对称合成。如果将根据该博士论文的这种在大肠杆菌中异源表达的酶用于在全细胞中合成，(S)-PAC的光学纯度为~43%ee。(S)-PAC的这个唯一的酶促不对称合成描述在由苯甲醛和乙醛或苯甲醛和丙酮酸起始的碳连接反应中。该反应由酶，即来自巴氏醋杆菌的丙酮酸脱羧酶的变体ApPDC-E469G催化，其中在位置号469中的谷氨酸被甘氨酸替换。在此通过分离的酶实现的最高对映体过量为89%，如由出版物RotherNeeGocke,Doerte;Kolter,Geraldine;Gerhards,Tina;Berthold,CatrineL.;Gauchenova,Ekaterina;Knoll,Michael;Pleiss,Juergen;Mueller,Michael;Schneider,Gunter;Pohl,Martina,ChemCatChem,2011,第3卷,10第1587-1596页描述。由现有技术出发，本发明的目的是提供不对称合成(S)-苯基乙酰基甲醇的替代性酶促方法，其实现(S)-苯基乙酰基甲醇的高对映体过量。在用全细胞制备时，对映体过量应大于43%。此外，不应形成副产物和区域异构体。在此，应能够使用非手性的廉价反应物。应实现(S)-苯基乙酰基甲醇的不对称合成。应防止手性产物混合物的昂贵分离。应提供可以用于由苯甲醛和丙酮酸或乙醛制备(S)-苯基乙酰基甲醇的酶，以及编码该酶的DNA。此外应提供制备该酶的方法。应克服现有技术中存在的缺点和问题。应提供制备(S)-苯基乙酰基甲醇的方法，其甚至在使用粗细胞提取物（Rohzellextrakt）或全细胞时实现高对映体过量。应累积地克服现有技术的缺点。由权利要求1的上位概念和从属权利要求出发，所述目的根据本发明通过在权利要求的特征部分中给出的特征来实现。该目的根据本发明通过裂解酶ApPDC-E469G的变体来实现，在该变体的位置号468中的异亮氨酸被具有相对于异亮氨酸减小的占据空间的氨基酸替换。此外，通过根据从属权利要求将裂解酶用于使苯甲醛与丙酮酸或乙醛反应成(S)-苯基乙酰基甲醇，实现该目的。通过本发明的裂解酶以及(S)-苯基乙酰基甲醇的制备方法，现在可以在使用全细胞时以93%ee和在使用粗细胞提取物时以85%ee制备(S)-苯基乙酰基甲醇。不形成副产物，特别是不产生区域异构体。由于该合成用非手性反应物也可实现，其是廉价的。可以免除对映体的分离。在制备(S)-苯基乙酰基甲醇时，也可以通过粗细胞提取物或全细胞实现高对映体过量。累积地消除根据现有技术描述的所有缺点。本发明的有利的扩展实施方案在从属权利要求中给出。根据本发明提供裂解酶，其中在相对于巴氏醋杆菌野生型修饰的蛋白ApPDC-E469G中在位置号468中的异亮氨酸被比异亮氨酸较不占空间的氨基酸置换。该裂解酶对于在制备(S)-苯基乙酰基甲醇时提高立体选择性而言具有有利影响。优选地，可以提及满足这些要求的下列裂解酶：-根据序列号1在位置号468中具有甘氨酸的ApPDC-E469G-I468G-根据序列号2在位置号468中具有丙氨酸的ApPDC-E469G-I468A-根据序列号3在位置号468中具有缬氨酸的ApPDC-E469G-I468V。为了改进对映体过量，也可以在制备(S)-苯基乙酰基甲醇时使用酶ApPDC-E469G-I468L、ApPDC-E469G-I468T、ApPDC-E469G-I468C或ApPDC-E469G-I468S。此外，提供编码所述酶的本发明的脱氧核糖核酸。根据本发明，这是编码酶ApPDC-E469G的变体的脱氧核糖核酸，其在位置号1402至1404中编码相对于异亮氨酸具有减小的占据空间的氨基酸。对于在所述位置中编码氨基酸甘氨酸的根据序列号1的实例，可以在位置号1402-1404中例如存在核酸GGT。例如，描述了根据序列号4的编码根据序列号1的在位置号468中具有甘氨酸的酶ApPDC-E469G-I468G的脱氧核糖核酸。对于本发明的根据序列号2的在位置号468中具有丙氨酸的酶ApPDC-E469G-I468A和根据序列号3的在位置号468中具有缬氨酸的酶ApPDC-E469G-I468V，可以通过替换位置号1402-1404中的相应核苷酸提供编码其的脱氧核糖核酸。编码本发明的酶的DNA可以根据本领域技术人员已知的方法通过定向或不定向的诱变来产生。在此，优选的是定向诱变。这些方法是本领域技术人员已知的。在具体的描述部分中，具体公开本发明的一个实施方案的制备实例。这种方法方式原则上也可以用于所有其它公开的脱氧核糖核酸和酶，以使得可以以类似途径制备所有的本发明的酶和脱氧核糖核酸。所述脱氧核糖核酸可以连接到载体，优选质粒中。作为空载体，可以例如使用pET-20b(+)、pET-21a-d(+)、pET-22b(+)、pET-23a-d(+)、pET-24a-d(+)、pET-25b(+)、pET-26b(+)、pET-27b(+)、pET-28a(+)、pET-29a-c(+)、pET-30a-c(+)、pET-31b(+)、pET-34b(+)、pET-35b(+)、pET-36b(+)、pET-37b(+)、pET-38b(+)，向其中连接相应的本发明DNA。替代地，也可以将该脱氧核糖核酸连接到基因组中。连接的脱氧核糖核酸是DNA序列，其编码酶ApPDC-E469G的变体并在位置号1402-1404中编码具有相对于异亮氨酸减小的占据空间的氨基酸。这些脱氧核糖核酸也可以编码酶ApPDC-E469G-I468L、ApPDC-E469G-I468T、ApPDC-E469G-I468C或ApPDC-E469G-I468S。优选地，连接的脱氧核糖核酸编码根据序列1、2和3的蛋白。根据本发明可以提供载体，其含有编码酶ApPDC-E469G的变体并在位置号1402-1404中编码具有相对于异亮氨酸减小或相等的占据空间的氨基酸的脱氧核糖核酸。优选地，该载体含有根据序列号4的脱氧核糖核酸。优选地，该载体是质粒。图1展示了本发明的质粒（pET-21a(+)载体卡）。在序列号5中例如示出本发明质粒的DNA序列，其含有根据序列号4的DNA。根据本发明，借助在位置号468中具有相对于异亮氨酸具有减小的占据空间的氨基酸的酶ApPDC-E469G的变体，优选选自序列号1、2或3的酶，使苯甲醛与丙酮酸或与乙醛根据式(1)反应成(S)-苯基乙酰基甲醇。式(1)该反应优选在水溶液中进行。pH值为5-9，优选6.5-8，特别优选6.5-7。对此，可以使用例如磷酸钾缓冲液、HEPES、MOPS、TEA或TRIS-HCl作为缓冲液。此外，可以使用磷酸硫胺素和硫酸镁作为辅因子。该反应可以在体内或在体外进行。对于在体内生产(S)-苯基乙酰基甲醇，可以使用例如大肠杆菌、棒状杆菌例如谷氨酸棒状杆菌或酵母例如酿酒酵母作为生产生物体。为此，该生产生物体通过本发明的DNA或包含该DNA的载体进行转化。该DNA也可以在生产生物体中引入基因组中。所用的基因在此异源表达。对于在体外生产，可以使用该生产生物体的分离的酶或细胞提取物。典型温度为20℃至40℃，优选20℃至30℃，特别优选为20℃至25℃的温度。反应时间可以为2h-48h，优选6h-24h，特别优选12h。下面介绍若干实施例，其不解释为具有限制性。这些反应可以以经典的配方（Ansatz）在反应烧瓶中在搅拌下进行。为了能够以高对映体过量制备(S)-PAC，通过诱变产生酶ApPDC-E469G的变体。变体ApPDC-E469G-I468G通过使用粗细胞提取物酶以85%ee生产(S)-PAC。通过使用在大肠杆菌中异源表达并用作廉价全细胞催化剂的ApPDC-E469G-I468G，可以以93%ee生成(S)-PAC。实施例1：20mM苯甲醛、400mM丙酮酸、2.5mM硫酸镁、100µM二磷酸硫胺素、20mg/mL（湿重）ApPDC-E469G-I468G（大肠杆菌的全细胞，其中ApPDC-E469G-I468G以表达形式存在）、50mM磷酸钾缓冲液pH6.5，25℃，反应时间：48h。(S)-PAC的对映体纯度：ee93%收率：67%。实施例2：20mM苯甲醛、400mM丙酮酸、2.5mM硫酸镁、100µM二磷酸硫胺素、1mg/mLApPDC-E469G-I468G（大肠杆菌的粗细胞提取物，其中ApPCD-E469G-I468G以表达形式存在）、50mM磷酸钾缓冲液pH6.5，25℃，反应时间：48h。(S)-PAC的对映体纯度：ee85%。下面示例性阐述本发明的酶和编码其的脱氧核糖核酸的制备。所示方法原则上也可以类似地用于制备其它的本发明裂解酶和编码其的脱氧核糖核酸。酶变体ApPDC-E469G-I468G的DNA的制备“位点饱和诱变”根据Reetz等人（Reetz,Kahakeaw等人2008）的变体的所谓的“位点饱和诱变”的方法由基因序列ApPDC-E469G（模板DNA）起始进行，以获得在位置号I468上的氨基酸替换。在这一方法中，使用编码20种天然氨基酸中的12种的NDT-密码子。聚合酶链反应（PCR）在最初的步骤中，借助聚合酶链反应（PCR）复制模板DNA，并在此同时通过使用所谓的简并引物（degeneriertenPrimer）和NDT密码子插入（eingefügt）突变。所用的引物从“EurofinsMWGOperon”公司订购（http://www.eurofinsgenomics.eu）并具有下列序列：用于制备ApPDC-E469G-I468NDT的位点饱和诱变的引物正向：5´CCGTGGCTATGTCNDTGGCATCGCCATTC3´（序列号6）反向：5´GAATGGCGATGCCAHNGACATAGCCACGG3´（序列号7）。首先制备母液并随后分为四个各为50µl的配制品。为了开始该反应，添加1µl“KOD热启动聚合酶”。PCR-反应配方：该反应在下列条件下进行：为了消化模板DNA，将1µl的酶DpnI（Eppendorf）添加到溶液中并在37℃下培养1h。整个配制品然后用“DNA纯化试剂盒”（化学品列表）在进一步转化前进行提纯。大肠杆菌BL21-DE3和大肠杆菌DH5α的转化菌株大肠杆菌BL21-DE3和大肠杆菌DH5α用通过“位点饱和诱变”制备的DNA进行转化。为此，向50µl感受态细胞中添加100ngDNA并在冰上培养30分钟。随后在42℃下进行热休克90秒。3分钟后，向冰上添加500µlSOC培养基并然后将该溶液在350转/分钟下且在37℃下在Eppendorf恒温混匀仪中培养45分钟。在完成培养后，将细胞悬浮液在13000转/分钟下在Eppendorf台式离心机中离心分离30秒，并将沉淀物（Pellet）接着再悬浮在100µl上清液中。浓缩至100µl的细胞悬浮液在LB琼脂平板（具有100µg/ml氨苄青霉素）上铺板并在37℃下倒立着培养16h。酶变体的表达转化的46个单菌落从板上用牙签挖出并在20℃和850转/分钟下在48孔Nerbe板（NerbePlusGmbH）的各具有1mlLB培养基的孔（“母板”）中分别培养24h。另一个孔用大肠杆菌BL21-DE3细胞接种，其事前与ApPDC-E469G-模板DNA类似地转化。在完成培养后，在48孔FlowerPlates®（m2p-labs,德国）中向各1.5ml自动诱导培养基中加入10µl细胞悬浮液。该FlowerPlate在20℃和850转/分钟下培养48h。向“母板”的剩余体积（990µl）加入300µl甘油并在-80℃下储存。细胞分解和碳连接将FlowerPlates®中表达的变体通过冷冻（48h，4℃）分解。在重新解冻后，将各500µl的细胞悬浮液转移到96孔板的两个孔中（双重测定）。将该板在4000转/分钟下离心3分钟并将沉淀物再悬浮到具有1mg/ml溶菌酶的420µlKpi缓冲液中。该板在20℃和400转/分钟下培养1h并随后重新在4000转/分钟下离心10分钟。将各250µl上清液移液到2mlNerbe板的各一个孔中并添加由40mM苯甲醛、400mM丙酮酸、4mM硫酸镁和400µM二磷酸硫胺素形成的250µl反应溶液。该板重新培养24h并随后分析反应溶液（参见HPLC分析）。HPLC分析向200µl碳连接反应溶液加入各200µl庚烷，涡旋并随后将各150µl的上部相转移到HPLC小瓶中。样品分析通过ChiralpakIC-3柱（ChiralTechnologiesInc.）在使用下列方法的情况下进行。HPLC程序典型的保留时间和用于定量的波长DNA分离和通过DNA测序的最佳酶变体的识别将碳连接反应中对于(S)-PAC提供最高ee值的酶的DNA由母板起始进行测序以识别突变。为此，首先将细胞用接种环从加入有甘油的母板转移到50mLLB培养基（+50µg/ml氨苄青霉素）中并在250mL锥形瓶中在37℃下培养。在12h培养后，将20mL细胞悬浮液离心分离（4000转/分钟，5min，4℃）。沉淀物中的细胞的DNA类似于制造商说明（QiagenN.V.）根据“QIAprep®SpinMiniprepKit”方法进行分离。为此，将DNA的浓度设定为100ng/µl并通过LGCGenomicsGmbH公司测序。LB（溶菌肉汤）培养基借助QuikChange®制备变体ApPDC-E469G-I468G的替代的定向方法制备酶变体ApPDC-E469G/I468G的另一方法例如是所谓的QuikChange®-PCR法（U.S.专利号5.789.166、5.932.419、6.391.548）。在PCR的这一变体的情况中，使用在待替换的DNA三联体密码的位置上带有相应的序列修饰的引物对。为了制备酶变体ApPDC-E469G/I468G，可以使用编码变体ApPDC-E469G的基因。这一所谓的DNA模板应在载体（例如pET22a）中以克隆形式存在。代替在位置I468中编码氨基酸色氨酸的三联体密码，必须使用在该位置上带有编码甘氨酸的突变的引物（即GGA、GGT、GGC或GGG）。这种QuikChange®-PCR法的所有其它参数以及所需引物的选择可以类似于制造商说明（AgilentTechnologies,Inc.）借助“QuikChange®定点诱变试剂盒”的指导来进行。用于制备变体ApPDC-E469G-I468G的QuikChange®-PCR法的DNA模板（ApPDC-E469G）。在序列表中，该序列以序列号8示出。序列号8在此例如对于编码根据序列号53的待修饰的蛋白ApPDC-E469G的DNA公开。但是，根据本发明，编码待修饰的起始蛋白的所有其它脱氧核糖核酸可以用于制备待修饰的酶。为其编码的核苷酸是本领域技术人员已知的。“全细胞”形式的变体的制备为了以1L的规模在全细胞中表达酶，首先将细胞从加入有甘油的母板用接种环转移到50mLLB培养基（+100µg/ml氨苄青霉素）中并在250mL锥形瓶中在120转/分钟和37℃下培养。在16h培养后，向1L自动诱导培养基中加入10mL培养物并在5L锥形瓶中在90转/分钟和20℃下培养72h。随后通过离心（4℃，6000转/分钟，30min）收获该细胞并在-20℃下储存以进一步使用。自动诱导培养基12g/l蛋白胨24g/l酵母提取物90mM磷酸钾缓冲液(pH7.5)0.5g/l葡萄糖2g/l乳糖0.01g/l氨苄青霉素6.3g/l甘油“细胞提取物”形式的变体的制备将10g以1L规模培养的细胞（参见““全细胞”形式的变体的制备”）用冷却到4℃的25mL破碎缓冲液（50mM磷酸钾pH6.5、100µM二磷酸硫胺素、2mM硫酸镁）在冰上再悬浮。随后，将再悬浮的细胞借助超声（SD14超声波发生器（HielscherUltrasonicsGmbH），4x2min超声处理，各1分钟来自冰的冷却）进行破碎。为了分离细胞碎片，将该溶液离心分离（45min，18000转/分钟，4℃）并将上清液（“细胞提取物”）转移到新容器中。“分离的酶”形式的变体的制备为了借助固定化金属离子亲合色谱法和尺寸排阻色谱法提纯ApPDC变体，使用“AmershamBioscience”公司的“ÄKTA™purifier”，以尤其检测蛋白-UV吸收（280nm）以及导电性并调节流速。为了提纯，将25mL制备的细胞提取物（参见““细胞提取物”形式的变体的制备”）以3mL/min的流速施加到体积为60mL的柱Ni-NTA-Superflow（QiagenN.V.）上，其事前用180mL上样缓冲液进行平衡。随后，继续用上样缓冲液以5ml/min的流速冲洗，以除去不结合或非常弱地结合在柱材料上的蛋白。在UV吸收（280nm）重新达到稳定基线后，漂洗缓冲液（50mM磷酸钾pH6.5、100µM二磷酸硫胺素、2mM硫酸镁、50mM咪唑）以5mL/min流速用于洗脱弱结合在柱材料上的蛋白。在重新达到稳定UV吸收（280nm）后，以5mL/min的流速使用洗脱缓冲液（50mM磷酸钾pH6.5、100µM二磷酸硫胺素、2mM硫酸镁、250mM咪唑）以洗脱目标蛋白。将IMAC的洗出液以10mL/min的流速施加到尺寸排阻色谱柱（1L柱体积，Sephadex-G25，GE-Health-care）上以进行再缓冲，该柱事先用2L再缓冲缓冲液（10mM磷酸钾pH6.5、100µM二磷酸硫胺素、2mM硫酸镁）冲洗。合并具有提高的UV吸收（280nm）的级分并在结晶皿中冷冻（-20℃）。为了冻干，向冷冻的蛋白溶液施加0.22mBar的负压3天。在本发明的另一个有利配置中，本发明的酶除了在位置号468中的根据本发明的替换之外，氨基酸色氨酸还在位置号543中被比色氨酸具有更小的占据空间的氨基酸替换。这些裂解酶此外还对在制备(S)-苯基乙酰基甲醇时提高立体选择性具有有利影响。通过本发明的这一优选的实施方案，可以实现98%ee的对映体过量。优选地，可以提及满足这些要求的下列裂解酶：-根据序列号9在位置号543中具有组氨酸的ApPCD-E469G-I468G-W543H-根据序列号10在位置号543中具有苯丙氨酸的ApPCD-E469G-I468G-W543F-根据序列号11在位置号543中具有脯氨酸的ApPCD-E469G-I468G-W543P-根据序列号12在位置号543中具有异亮氨酸的ApPCD-E469G-I468G-W543I-根据序列号13在位置号543中具有亮氨酸的ApPCD-E469G-I468G-W543L-根据序列号14在位置号543中具有甲硫氨酸的ApPCD-E469G-I468G-W543M-根据序列号15在位置号543中具有缬氨酸的ApPCD-E469G-I468G-W543V-根据序列号16在位置号543中具有丙氨酸的ApPCD-E469G-I468G-W543A-根据序列号17在位置号543中具有酪氨酸的ApPCD-E469G-I468G-W543Y-根据序列号18在位置号543中具有苏氨酸的ApPCD-E469G-I468G-W543T-根据序列号19在位置号543中具有甘氨酸的ApPCD-E469G-I468G-W543G-根据序列号20在位置号543中具有丝氨酸的ApPCD-E469G-I468G-W543S-根据序列号21在位置号543中具有半胱氨酸ApPCD-E469G-I468G-W543C以及-根据序列号22在位置号543中具有组氨酸的ApPCD-E469G-I468A-W543H-根据序列号23在位置号543中具有苯丙氨酸的ApPCD-E469G-I468A-W543F-根据序列号24在位置号543中具有脯氨酸的ApPCD-E469G-I468A-W543P-根据序列号25在位置号543中具有异亮氨酸的ApPCD-E469G-I468A-W543I-根据序列号26在位置号543中具有亮氨酸的ApPCD-E469G-I468A-W543L-根据序列号27在位置号543中具有甲硫氨酸的ApPCD-E469G-I468A-W543M-根据序列号28在位置号543中具有缬氨酸的ApPCD-E469G-I468A-W543V-根据序列号29在位置号543中具有丙氨酸的ApPCD-E469G-I468A-W543A-根据序列号30在位置号543中具有酪氨酸的ApPCD-E469G-I468A-W543Y-根据序列号31在位置号543中具有苏氨酸的ApPCD-E469G-I468A-W543T-根据序列号32在位置号543中具有甘氨酸的ApPCD-E469G-I468A-W543G-根据序列号33在位置号543中具有丝氨酸的ApPCD-E469G-I468A-W543S-根据序列号34在位置号543中具有半胱氨酸的ApPCD-E469G-I468A-W543C和-根据序列号35在位置号543中具有组氨酸的ApPCD-E469G-I468V-W543H-根据序列号36在位置号543中具有苯丙氨酸的ApPCD-E469G-I468V-W543F-根据序列号37在位置号543中具有脯氨酸的ApPCD-E469G-I468V-W543P-根据序列号38在位置号543中具有异亮氨酸的ApPCD-E469G-I468V-W543I-根据序列号39在位置号543中具有亮氨酸的ApPCD-E469G-I468V-W543L-根据序列号40在位置号543中具有甲硫氨酸的ApPCD-E469G-I468V-W543M-根据序列号41在位置号543中具有缬氨酸的ApPCD-E469G-I468V-W543V-根据序列号42在位置号543中具有丙氨酸的ApPCD-E469G-I468V-W543A-根据序列号43在位置号543中具有酪氨酸的ApPCD-E469G-I468V-W543Y-根据序列号44在位置号543中具有苏氨酸的ApPCD-E469G-I468V-W543T-根据序列号45在位置号543中具有甘氨酸的ApPCD-E469G-I468V-W543G-根据序列号46在位置号543中具有丝氨酸的ApPCD-E469G-I468V-W543S-根据序列号47在位置号543中具有半胱氨酸的ApPCD-E469G-I468V-W543C。此外，根据本发明提供编码所述酶的脱氧核糖核酸。根据本发明，这是编码酶ApPDC-E469G的变体并在位置号1402-1404中编码具有相对于异亮氨酸减小的占据空间的氨基酸且此外在位置号1627-1629中编码比色氨酸较小占据空间的氨基酸的脱氧核糖核酸。对于根据序列号23的蛋白的实例，可以在相应脱氧核糖核酸的位置号1402-1404中例如存在核酸GCC并在位置号1627-1629中存在核酸TTT。相应的核苷酸序列在序列表中列在编号48下。编码根据序列9至47的本发明酶的DNA可以根据本领域技术人员已知的方法通过定向或不定向的诱变来产生。在此，优选的是定向诱变。这些方法是本领域技术人员已知的。在具体的描述部分中，具体公开本发明的一个实施方案的制备实例。这种方法方式原则上也可以用于所有其它公开的脱氧核糖核酸和酶，以使得可以以类似途径制备所有的本发明的酶和脱氧核糖核酸。所述脱氧核糖核酸可以连接到载体，优选质粒中。作为空载体，可以例如使用pET-20b(+)、pET-21a-d(+)、pET-22b(+)、pET-23a-d(+)、pET-24a-d(+)、pET-25b(+)、pET-26b(+)、pET-27b(+)、pET-28a(+)、pET-29a-c(+)、pET-30a-c(+)、pET-31b(+)、pET-34b(+)、pET-35b(+)、pET-36b(+)、pET-37b(+)、pET-38b(+)，向其中连接相应的本发明DNA。替代地，也可以将该脱氧核糖核酸连接到基因组中。连接的脱氧核糖核酸是DNA，其编码酶ApPDC-E469G的变体并在位置号1402-1404中编码具有相对于异亮氨酸减小的占据空间的氨基酸且在位置号1627-1629中编码具有相对于色氨酸减小的占据空间的氨基酸。优选地，连接的脱氧核糖核酸编码根据序列9至47的蛋白。根据本发明可以提供载体，其含有编码酶ApPDC-E469G的变体并在位置号1402-1404中编码具有相对于异亮氨酸减小的占据空间的氨基酸且在位置号1627-1629中编码具有相对于色氨酸减小的占据空间的氨基酸的脱氧核糖核酸。优选地，该载体含有根据序列号48的脱氧核糖核酸。优选地，该载体是质粒。在序列号49中例如示出本发明质粒的序列，其含有根据序列号48的DNA。根据本发明，还在本发明的有利配置中借助在位置号468中存在具有相对于异亮氨酸减小的占据空间的氨基酸且在位置号543中存在具有相对于色氨酸减小的占据空间的氨基酸的酶ApPDC-E469G的变体，优选选自根据序列号9至47的酶，使苯甲醛与丙酮酸或与乙醛根据式(1)反应成(S)-苯基乙酰基甲醇。式(1)该反应优选在水溶液中进行。pH值为5-9，优选6.5-8，特别优选6.5-7。对此，可以使用例如磷酸钾缓冲液、HEPES、MOPS、TEA或TRIS-HCl作为缓冲液。此外，可以使用磷酸硫胺素和硫酸镁作为辅因子。该反应可以在体内或在体外进行。对于在体内生产(S)-苯基乙酰基甲醇，可以使用例如大肠杆菌、棒状杆菌例如谷氨酸棒状杆菌或酵母例如酿酒酵母作为生产生物体。为此，该生产生物体通过本发明的DNA或包含该DNA的载体进行转化。该DNA也可以在生产生物体中引入基因组中。所用的基因在此异源表达。对于在体外生产，可以使用该生产生物体的分离的酶或细胞提取物。典型温度为20℃至40℃，优选20℃至30℃，特别优选为20℃至25℃的温度。反应时间可以为2h-48h，优选6h-24h，特别优选12h。下面介绍若干实施例，其不解释为具有限制性。这些反应可以以经典的配方在反应烧瓶中在搅拌下进行。为了能够以高对映体过量制备(S)-PAC，通过诱变产生酶ApPDC-E469G的变体。变体ApPDC-E469G-I468A-W543F通过使用分离的酶以98%ee生产(S)-PAC。通过使用在大肠杆菌中异源表达并用作廉价全细胞催化剂的ApPDC-E469G-I468A-W543F，可以以96%ee生成(S)-PAC。实施例3：20mM苯甲醛、400mM丙酮酸、2.5mM硫酸镁、100µM二磷酸硫胺素、20mg/mL（湿重）ApPDC-E469G-I468A-W543F（大肠杆菌的全细胞，其中ApPDC-E469G-I468A-W543F以表达形式存在）、50mM磷酸钾缓冲液pH6.5，25℃，反应时间：48h。(S)-PAC的对映体纯度：ee96%收率：73%。实施例4：20mM苯甲醛、400mM丙酮酸、2.5mM硫酸镁、100µM二磷酸硫胺素、1mg/mLApPDC-E469G-I468A-W543F20mM苯甲醛、400mM丙酮酸、2.5mM硫酸镁、100µM二磷酸硫胺素、1mg/mLApPDC-E469G-I468G（分离的酶）、50mM磷酸钾缓冲液pH6.5，25℃，反应时间：48h。，50mM磷酸钾缓冲液pH6.5，25℃，反应时间：48h。(S)-PAC的对映体纯度：ee98%收率：45%。借助QuikChange®由ApPDC-E469G-I468A-W543起始制备变体ApPDC-E469G-I468A-W543F的定向方法为了制备酶变体ApPDC-E469G-I468A-W543F，使用所谓的QuikChange®-PCR法（U.S.专利号5.789.166、5.932.419、6.391.548）。在PCR的这一变体的情况中，使用在待替换的DNA三联体密码的位置上带有相应的序列修饰的引物对。为了制备酶变体ApPDC-E469G-I468A-W543F，使用编码变体ApPDC-E469G-I468A-W543的基因。这一所谓的DNA模板在载体（例如pET22a）中以克隆形式存在。代替在位置W543中编码氨基酸色氨酸的三联体密码，必须使用在该位置上带有编码苯丙氨酸的突变的引物（即例如TTC或TTT）。用于由ApPDC-E469G-I468A-W543起始制备酶变体ApPDC-E469G-I468A-W543F的QuikChange®-PCR法的引物正向：5´accttgcggccgaattgaaccagcatatccgtgc3´（序列号50）反向：5´gcacggatatgctggttcaattcggccgcaaggt3´（序列号51）。首先制备母液并随后分为四个各为50µl的配制品。为了开始该反应，添加1µl“PfuTurbo®DNA聚合酶”。PCR-反应配方：该反应在下列条件下进行：为了消化模板DNA，将1µl的酶DpnI（Eppendorf）添加到溶液中并在37℃下培养1h。整个配制品然后用“DNA纯化试剂盒”（化学品列表）在进一步转化前进行提纯。这一QuikChange®-PCR法的所有其它参数以及对于其它酶变体的所需引物的选择可以类似于制造商（AgilentTechnologies,Inc.）的说明借助“QuikChange®定点诱变试剂盒”的指导来使用。制备变体ApPDC-E469G-I468A-W543F的QuikChange®-PCR法的DNA模板（ApPDC-E469G-I468A）。序列号52在此例如对于编码根据序列号2的待修饰的蛋白ApPDC-E469G-I468A的DNA公开。但是，根据本发明，编码待修饰的起始蛋白的所有其它脱氧核糖核酸可以用于制备待修饰的酶。为其编码的核苷酸是本领域技术人员已知的。大肠杆菌BL21-DE3和大肠杆菌DH5α的转化菌株大肠杆菌BL21-DE3和大肠杆菌DH5α用通过“位点饱和诱变”制备的DNA进行转化。为此，向50µl感受态细胞中添加100ngDNA并在冰上培养30分钟。随后在42℃下进行热休克90秒。3分钟后，向冰上添加500µlSOC培养基并然后将该溶液在350转/分钟下且在37℃下在Eppendorf恒温混匀仪中培养45分钟。在完成培养后，将细胞悬浮液在13000转/分钟下在Eppendorf台式离心机中离心分离30秒，并将沉淀物接着再悬浮在100µl上清液中。浓缩至100µl的细胞悬浮液在LB琼脂平板（具有100µg/ml氨苄青霉素）上铺板并在37℃下倒立着培养16h。DNA分离和通过DNA测序的最佳酶变体的识别将碳连接反应中对于(S)-PAC提供最高ee值的酶的DNA由母板起始进行测序以识别突变。为此，首先将细胞用接种环从加入有甘油的母板转移到50mLLB培养基（+50µg/ml氨苄青霉素）中并在250mL锥形瓶中在37℃下培养。在12h培养后，将20mL细胞悬浮液离心分离（4000转/分钟，5min，4℃）。沉淀物中的细胞的DNA类似于制造商说明（QiagenN.V.）根据“QIAprep®SpinMiniprepKit”方法进行分离。为此，将DNA的浓度设定为100ng/µl并通过LGCGenomicsGmbH公司测序。LB（溶菌肉汤）培养基“全细胞”形式的变体的制备为了以1L的规模在全细胞中表达酶，首先将细胞从加入有甘油的母板用接种环转移到50mLLB培养基（+100µg/ml氨苄青霉素）中并在250mL锥形瓶中在120转/分钟和37℃下培养。在16h培养后，向1L自动诱导培养基中加入10mL培养物并在5L锥形瓶中在90转/分钟和20℃下培养72h。随后通过离心（4℃，6000转/分钟，30min）收获该细胞并在-20℃下储存以进一步使用。自动诱导培养基12g/l蛋白胨24g/l酵母提取物90mM磷酸钾缓冲液(pH7.5)0.5g/l葡萄糖2g/l乳糖0.01g/l氨苄青霉素6.3g/l甘油分离的酶的形式的变体的制备将10g以1L规模培养的细胞用冷却到4℃的25mL破碎缓冲液（50mM磷酸钾pH6.5、100µM二磷酸硫胺素、2mM硫酸镁）在冰上再悬浮。随后，将再悬浮的细胞借助超声（SD14超声波发生器（HielscherUltrasonicsGmbH），4x2min超声处理，各1分钟来自冰的冷却）进行破碎。为了分离细胞碎片，将该溶液离心分离（45min，18000转/分钟，4℃）并将上清液（“细胞提取物”）转移到新容器中。为了借助固定化金属离子亲合色谱法和尺寸排阻色谱法提纯ApPDC变体，使用“AmershamBioscience”公司的“ÄKTA™purifier”，以尤其检测蛋白-UV吸收（280nm）以及导电性并调节流速。为了提纯，将制备的细胞提取物（~25mL）以3mL/min的流速施加到体积为60mL的柱Ni-NTA-Superflow（QiagenN.V.）上，其事前用180mL上样缓冲液进行平衡。随后，继续用上样缓冲液以5ml/min的流速冲洗，以除去不结合或非常弱地结合在柱材料上的蛋白。在UV吸收（280nm）重新达到稳定基线后，漂洗缓冲液（50mM磷酸钾pH6.5、100µM二磷酸硫胺素、2mM硫酸镁、50mM咪唑）以5mL/min流速用于洗脱弱结合在柱材料上的蛋白。在重新达到稳定UV吸收（280nm）后，以5mL/min的流速使用洗脱缓冲液（50mM磷酸钾pH6.5、100µM二磷酸硫胺素、2mM硫酸镁、250mM咪唑）以洗脱目标蛋白。将IMAC的洗出液以10mL/min的流速施加到尺寸排阻色谱柱（1L柱体积,Sephadex-G25,GE-Healthcare）上以进行再缓冲，该柱事先用2L再缓冲缓冲液（10mM磷酸钾pH6.5、100µM二磷酸硫胺素、2mM硫酸镁）冲洗。合并具有提高的UV吸收（280nm）的级分并在结晶皿中冷冻（-20℃）。为了冻干，向冷冻的蛋白溶液施加0.22mBar的负压3天。所产生的粉末的蛋白含量为20%。纯度（目标蛋白相对于外源蛋白的含量）为>90%。序列表<110>ForschungszentrumJülichGmbH<120>裂解酶以及不对称合成苯基乙酰基甲醇的方法<130>PT1.2668<160>53<170>PatentInversion3.5<210>1<211>560<212>PRT<213>人工<220><223>人工制备<400>1MetThrTyrThrValGlyMetTyrLeuAlaGluArgLeuValGlnIle151015GlyLeuLysHisHisPheAlaValAlaGlyAspTyrAsnLeuValLeu202530LeuAspGlnLeuLeuLeuAsnLysAspMetLysGlnIleTyrCysCys354045AsnGluLeuAsnCysGlyPheSerAlaGluGlyTyrAlaArgSerAsn505560GlyAlaAlaAlaAlaValValThrPheSerValGlyAlaIleSerAla65707580MetAsnAlaLeuGlyGlyAlaTyrAlaGluAsnLeuProValIleLeu859095IleSerGlyAlaProAsnSerAsnAspGlnGlyThrGlyHisIleLeu100105110HisHisThrIleGlyLysThrAspTyrSerTyrGlnLeuGluMetAla115120125ArgGlnValThrCysAlaAlaGluSerIleThrAspAlaHisSerAla130135140ProAlaLysIleAspHisValIleArgThrAlaLeuArgGluArgLys145150155160ProAlaTyrLeuAspIleAlaCysAsnIleAlaSerGluProCysVal165170175ArgProGlyProValSerSerLeuLeuSerGluProGluIleAspHis180185190ThrSerLeuLysAlaAlaValAspAlaThrValAlaLeuLeuGluLys195200205SerAlaSerProValMetLeuLeuGlySerLysLeuArgAlaAlaAsn210215220AlaLeuAlaAlaThrGluThrLeuAlaAspLysLeuGlnCysAlaVal225230235240ThrIleMetAlaAlaAlaLysGlyPhePheProGluAspHisAlaGly245250255PheArgGlyLeuTyrTrpGlyGluValSerAsnProGlyValGlnGlu260265270LeuValGluThrSerAspAlaLeuLeuCysIleAlaProValPheAsn275280285AspTyrSerThrValGlyTrpSerAlaTrpProLysGlyProAsnVal290295300IleLeuAlaGluProAspArgValThrValAspGlyArgAlaTyrAsp305310315320GlyPheThrLeuArgAlaPheLeuGlnAlaLeuAlaGluLysAlaPro325330335AlaArgProAlaSerAlaGlnLysSerSerValProThrCysSerLeu340345350ThrAlaThrSerAspGluAlaGlyLeuThrAsnAspGluIleValArg355360365HisIleAsnAlaLeuLeuThrSerAsnThrThrLeuValAlaGluThr370375380GlyAspSerTrpPheAsnAlaMetArgMetThrLeuProArgGlyAla385390395400ArgValGluLeuGluMetGlnTrpGlyHisIleGlyTrpSerValPro405410415SerAlaPheGlyAsnAlaMetGlySerGlnAspArgGlnHisValVal420425430MetValGlyAspGlySerPheGlnLeuThrAlaGlnGluValAlaGln435440445MetValArgTyrGluLeuProValIleIlePheLeuIleAsnAsnArg450455460GlyTyrValGlyGlyIleAlaIleHisAspGlyProTyrAsnTyrIle465470475480LysAsnTrpAspTyrAlaGlyLeuMetGluValPheAsnAlaGlyGlu485490495GlyHisGlyLeuGlyLeuLysAlaThrThrProLysGluLeuThrGlu500505510AlaIleAlaArgAlaLysAlaAsnThrArgGlyProThrLeuIleGlu515520525CysGlnIleAspArgThrAspCysThrAspMetLeuValGlnTrpGly530535540ArgLysValAlaSerThrAsnAlaArgLysThrThrLeuAlaLeuGlu545550555560<210>2<211>560<212>PRT<213>人工<220><223>人工制备<400>2MetThrTyrThrValGlyMetTyrLeuAlaGluArgLeuValGlnIle151015GlyLeuLysHisHisPheAlaValAlaGlyAspTyrAsnLeuValLeu202530LeuAspGlnLeuLeuLeuAsnLysAspMetLysGlnIleTyrCysCys354045AsnGluLeuAsnCysGlyPheSerAlaGluGlyTyrAlaArgSerAsn505560GlyAlaAlaAlaAlaValValThrPheSerValGlyAlaIleSerAla65707580MetAsnAlaLeuGlyGlyAlaTyrAlaGluAsnLeuProValIleLeu859095IleSerGlyAlaProAsnSerAsnAspGlnGlyThrGlyHisIleLeu100105110HisHisThrIleGlyLysThrAspTyrSerTyrGlnLeuGluMetAla115120125ArgGlnValThrCysAlaAlaGluSerIleThrAspAlaHisSerAla130135140ProAlaLysIleAspHisValIleArgThrAlaLeuArgGluArgLys145150155160ProAlaTyrLeuAspIleAlaCysAsnIleAlaSerGluProCysVal165170175ArgProGlyProValSerSerLeuLeuSerGluProGluIleAspHis180185190ThrSerLeuLysAlaAlaValAspAlaThrValAlaLeuLeuGluLys195200205SerAlaSerProValMetLeuLeuGlySerLysLeuArgAlaAlaAsn210215220AlaLeuAlaAlaThrGluThrLeuAlaAspLysLeuGlnCysAlaVal225230235240ThrIleMetAlaAlaAlaLysGlyPhePheProGluAspHisAlaGly245250255PheArgGlyLeuTyrTrpGlyGluValSerAsnProGlyValGlnGlu260265270LeuValGluThrSerAspAlaLeuLeuCysIleAlaProValPheAsn275280285AspTyrSerThrValGlyTrpSerAlaTrpProLysGlyProAsnVal290295300IleLeuAlaGluProAspArgValThrValAspGlyArgAlaTyrAsp305310315320GlyPheThrLeuArgAlaPheLeuGlnAlaLeuAlaGluLysAlaPro325330335AlaArgProAlaSerAlaGlnLysSerSerValProThrCysSerLeu340345350ThrAlaThrSerAspGluAlaGlyLeuThrAsnAspGluIleValArg355360365HisIleAsnAlaLeuLeuThrSerAsnThrThrLeuValAlaGluThr370375380GlyAspSerTrpPheAsnAlaMetArgMetThrLeuProArgGlyAla385390395400ArgValGluLeuGluMetGlnTrpGlyHisIleGlyTrpSerValPro405410415SerAlaPheGlyAsnAlaMetGlySerGlnAspArgGlnHisValVal420425430MetValGlyAspGlySerPheGlnLeuThrAlaGlnGluValAlaGln435440445MetValArgTyrGluLeuProValIleIlePheLeuIleAsnAsnArg450455460GlyTyrValAlaGlyIleAlaIleHisAspGlyProTyrAsnTyrIle465470475480LysAsnTrpAspTyrAlaGlyLeuMetGluValPheAsnAlaGlyGlu485490495GlyHisGlyLeuGlyLeuLysAlaThrThrProLysGluLeuThrGlu500505510AlaIleAlaArgAlaLysAlaAsnThrArgGlyProThrLeuIleGlu515520525CysGlnIleAspArgThrAspCysThrAspMetLeuValGlnTrpGly530535540ArgLysValAlaSerThrAsnAlaArgLysThrThrLeuAlaLeuGlu545550555560<210>3<211>560<212>PRT<213>人工<220><223>人工制备<400>3MetThrTyrThrValGlyMetTyrLeuAlaGluArgLeuValGlnIle151015GlyLeuLysHisHisPheAlaValAlaGlyAspTyrAsnLeuValLeu202530LeuAspGlnLeuLeuLeuAsnLysAspMetLysGlnIleTyrCysCys354045AsnGluLeuAsnCysGlyPheSerAlaGluGlyTyrAlaArgSerAsn505560GlyAlaAlaAlaAlaValValThrPheSerValGlyAlaIleSerAla65707580MetAsnAlaLeuGlyGlyAlaTyrAlaGluAsnLeuProValIleLeu859095IleSerGlyAlaProAsnSerAsnAspGlnGlyThrGlyHisIleLeu100105110HisHisThrIleGlyLysThrAspTyrSerTyrGlnLeuGluMetAla115120125ArgGlnValThrCysAlaAlaGluSerIleThrAspAlaHisSerAla130135140ProAlaLysIleAspHisValIleArgThrAlaLeuArgGluArgLys145150155160ProAlaTyrLeuAspIleAlaCysAsnIleAlaSerGluProCysVal165170175ArgProGlyProValSerSerLeuLeuSerGluProGluIleAspHis180185190ThrSerLeuLysAlaAlaValAspAlaThrValAlaLeuLeuGluLys195200205SerAlaSerProValMetLeuLeuGlySerLysLeuArgAlaAlaAsn210215220AlaLeuAlaAlaThrGluThrLeuAlaAspLysLeuGlnCysAlaVal225230235240ThrIleMetAlaAlaAlaLysGlyPhePheProGluAspHisAlaGly245250255PheArgGlyLeuTyrTrpGlyGluValSerAsnProGlyValGlnGlu260265270LeuValGluThrSerAspAlaLeuLeuCysIleAlaProValPheAsn275280285AspTyrSerThrValGlyTrpSerAlaTrpProLysGlyProAsnVal290295300IleLeuAlaGluProAspArgValThrValAspGlyArgAlaTyrAsp305310315320GlyPheThrLeuArgAlaPheLeuGlnAlaLeuAlaGluLysAlaPro325330335AlaArgProAlaSerAlaGlnLysSerSerValProThrCysSerLeu340345350ThrAlaThrSerAspGluAlaGlyLeuThrAsnAspGluIleValArg355360365HisIleAsnAlaLeuLeuThrSerAsnThrThrLeuValAlaGluThr370375380GlyAspSerTrpPheAsnAlaMetArgMetThrLeuProArgGlyAla385390395400ArgValGluLeuGluMetGlnTrpGlyHisIleGlyTrpSerValPro405410415SerAlaPheGlyAsnAlaMetGlySerGlnAspArgGlnHisValVal420425430MetValGlyAspGlySerPheGlnLeuThrAlaGlnGluValAlaGln435440445MetValArgTyrGluLeuProValIleIlePheLeuIleAsnAsnArg450455460GlyTyrValValGlyIleAlaIleHisAspGlyProTyrAsnTyrIle465470475480LysAsnTrpAspTyrAlaGlyLeuMetGluValPheAsnAlaGlyGlu485490495GlyHisGlyLeuGlyLeuLysAlaThrThrProLysGluLeuThrGlu500505510AlaIleAlaArgAlaLysAlaAsnThrArgGlyProThrLeuIleGlu515520525CysGlnIleAspArgThrAspCysThrAspMetLeuValGlnTrpGly530535540ArgLysValAlaSerThrAsnAlaArgLysThrThrLeuAlaLeuGlu545550555560<210>4<211>1680<212>DNA<213>人工<220><223>人工制备<400>4atgacctatactgttggcatgtatcttgcagaacgccttgtacagatcgggctgaagcat60cacttcgccgtggcgggcgactacaatctcgttcttctggatcagttgctcctcaacaag120gacatgaaacagatctattgctgcaatgagttgaactgtggcttcagcgcggaaggctac180gcccgttctaacggggctgcggcagcggttgtcaccttcagcgttggcgccatttccgcc240atgaacgccctcggcggcgcctatgccgaaaacctgccggttatcctgatttccggcgcg300cccaacagcaatgatcagggcacaggtcatatcctgcatcacacaatcggcaagacggat360tacagctaccagcttgaaatggcccgtcaggtcacctgtgccgccgaaagcattaccgac420gctcactccgccccggccaagattgaccacgtcattcgcacggcgctgcgcgagcgtaag480ccggcctatctggacatcgcgtgcaacattgcctccgagccctgcgtgcggcctggccct540gtcagcagcctgctgtccgagcctgaaatcgaccacacgagcctgaaggccgcagtggac600gccacggttgccttgctggaaaaatcggccagccccgtcatgctgctgggcagcaagctg660cgggccgccaacgcactggccgcaaccgaaacgctggcagacaagctgcaatgcgcggtg720accatcatggcggccgcgaaaggctttttccccgaagaccacgcgggtttccgcggcctg780tactggggcgaagtctcgaaccccggcgtgcaggaactggtggagacctccgacgcactg840ctgtgcatcgcccccgtattcaacgactattcaacagtcggctggtcggcatggcccaag900ggccccaatgtgattctggctgagcccgaccgcgtaacggtcgatggccgcgcctatgac960ggctttaccctgcgcgccttcctgcaggctctggcggaaaaagcccccgcgcgcccggcc1020tccgcacagaaaagcagcgtcccgacgtgctcgctcaccgcgacatccgatgaagccggt1080ctgacgaatgacgaaatcgtccgtcatatcaacgccctgctgacatcaaacacgacgctg1140gtggcagaaaccggcgattcatggttcaatgccatgcgcatgaccctgccgcgcggtgcg1200cgcgtggaactggaaatgcagtggggccatatcggctggtccgtgccctccgccttcggc1260aatgccatgggctcgcaggaccgccagcatgtggtgatggtaggcgatggctccttccag1320cttaccgcgcaggaagtggctcagatggtgcgctacgaactgcccgtcattatctttctg1380atcaacaaccgtggctatgtcggtggcatcgccattcatgacggcccgtacaactatatc1440aagaactgggattacgccggcctgatggaagtcttcaacgccggagaaggccatggactt1500ggcctgaaagccaccaccccgaaggaactgacagaagccatcgccagggcaaaagccaat1560acccgcggcccgacgctgatcgaatgccagatcgaccgcacggactgcacggatatgctg1620gttcaatggggccgcaaggttgcctcaaccaacgcgcgcaagaccactctggccctcgag1680<210>5<211>7041<212>DNA<213>人工<220><223>人工制备<400>5tggcgaatgggacgcgccctgtagcggcgcattaagcgcggcgggtgtggtggttacgcg60cagcgtgaccgctacacttgccagcgccctagcgcccgctcctttcgctttcttcccttc120ctttctcgccacgttcgccggctttccccgtcaagctctaaatcgggggctccctttagg180gttccgatttagtgctttacggcacctcgaccccaaaaaacttgattagggtgatggttc240acgtagtgggccatcgccctgatagacggtttttcgccctttgacgttggagtccacgtt300ctttaatagtggactcttgttccaaactggaacaacactcaaccctatctcggtctattc360ttttgatttataagggattttgccgatttcggcctattggttaaaaaatgagctgattta420acaaaaatttaacgcgaattttaacaaaatattaacgtttacaatttcaggtggcacttt480tcggggaaatgtgcgcggaacccctatttgtttatttttctaaatacattcaaatatgta540tccgctcatgagacaataaccctgataaatgcttcaataatattgaaaaaggaagagtat600gagtattcaacatttccgtgtcgcccttattcccttttttgcggcattttgccttcctgt660ttttgctcacccagaaacgctggtgaaagtaaaagatgctgaagatcagttgggtgcacg720agtgggttacatcgaactggatctcaacagcggtaagatccttgagagttttcgccccga780agaacgttttccaatgatgagcacttttaaagttctgctatgtggcgcggtattatcccg840tattgacgccgggcaagagcaactcggtcgccgcatacactattctcagaatgacttggt900tgagtactcaccagtcacagaaaagcatcttacggatggcatgacagtaagagaattatg960cagtgctgccataaccatgagtgataacactgcggccaacttacttctgacaacgatcgg1020aggaccgaaggagctaaccgcttttttgcacaacatgggggatcatgtaactcgccttga1080tcgttgggaaccggagctgaatgaagccataccaaacgacgagcgtgacaccacgatgcc1140tgcagcaatggcaacaacgttgcgcaaactattaactggcgaactacttactctagcttc1200ccggcaacaattaatagactggatggaggcggataaagttgcaggaccacttctgcgctc1260ggcccttccggctggctggtttattgctgataaatctggagccggtgagcgtgggtctcg1320cggtatcattgcagcactggggccagatggtaagccctcccgtatcgtagttatctacac1380gacggggagtcaggcaactatggatgaacgaaatagacagatcgctgagataggtgcctc1440actgattaagcattggtaactgtcagaccaagtttactcatatatactttagattgattt1500aaaacttcatttttaatttaaaaggatctaggtgaagatcctttttgataatctcatgac1560caaaatcccttaacgtgagttttcgttccactgagcgtcagaccccgtagaaaagatcaa1620aggatcttcttgagatcctttttttctgcgcgtaatctgctgcttgcaaacaaaaaaacc1680accgctaccagcggtggtttgtttgccggatcaagagctaccaactctttttccgaaggt1740aactggcttcagcagagcgcagataccaaatactgtccttctagtgtagccgtagttagg1800ccaccacttcaagaactctgtagcaccgcctacatacctcgctctgctaatcctgttacc1860agtggctgctgccagtggcgataagtcgtgtcttaccgggttggactcaagacgatagtt1920accggataaggcgcagcggtcgggctgaacggggggttcgtgcacacagcccagcttgga1980gcgaacgacctacaccgaactgagatacctacagcgtgagctatgagaaagcgccacgct2040tcccgaagggagaaaggcggacaggtatccggtaagcggcagggtcggaacaggagagcg2100cacgagggagcttccagggggaaacgcctggtatctttatagtcctgtcgggtttcgcca2160cctctgacttgagcgtcgatttttgtgatgctcgtcaggggggcggagcctatggaaaaa2220cgccagcaacgcggcctttttacggttcctggccttttgctggccttttgctcacatgtt2280ctttcctgcgttatcccctgattctgtggataaccgtattaccgcctttgagtgagctga2340taccgctcgccgcagccgaacgaccgagcgcagcgagtcagtgagcgaggaagcggaaga2400gcgcctgatgcggtattttctccttacgcatctgtgcggtatttcacaccgcatatatgg2460tgcactctcagtacaatctgctctgatgccgcatagttaagccagtatacactccgctat2520cgctacgtgactgggtcatggctgcgccccgacacccgccaacacccgctgacgcgccct2580gacgggcttgtctgctcccggcatccgcttacagacaagctgtgaccgtctccgggagct2640gcatgtgtcagaggttttcaccgtcatcaccgaaacgcgcgaggcagctgcggtaaagct2700catcagcgtggtcgtgaagcgattcacagatgtctgcctgttcatccgcgtccagctcgt2760tgagtttctccagaagcgttaatgtctggcttctgataaagcgggccatgttaagggcgg2820ttttttcctgtttggtcactgatgcctccgtgtaagggggatttctgttcatgggggtaa2880tgataccgatgaaacgagagaggatgctcacgatacgggttactgatgatgaacatgccc2940ggttactggaacgttgtgagggtaaacaactggcggtatggatgcggcgggaccagagaa3000aaatcactcagggtcaatgccagcgcttcgttaatacagatgtaggtgttccacagggta3060gccagcagcatcctgcgatgcagatccggaacataatggtgcagggcgctgacttccgcg3120tttccagactttacgaaacacggaaaccgaagaccattcatgttgttgctcaggtcgcag3180acgttttgcagcagcagtcgcttcacgttcgctcgcgtatcggtgattcattctgctaac3240cagtaaggcaaccccgccagcctagccgggtcctcaacgacaggagcacgatcatgcgca3300cccgtggggccgccatgccggcgataatggcctgcttctcgccgaaacgtttggtggcgg3360gaccagtgacgaaggcttgagcgagggcgtgcaagattccgaataccgcaagcgacaggc3420cgatcatcgtcgcgctccagcgaaagcggtcctcgccgaaaatgacccagagcgctgccg3480gcacctgtcctacgagttgcatgataaagaagacagtcataagtgcggcgacgatagtca3540tgccccgcgcccaccggaaggagctgactgggttgaaggctctcaagggcatcggtcgag3600atcccggtgcctaatgagtgagctaacttacattaattgcgttgcgctcactgcccgctt3660tccagtcgggaaacctgtcgtgccagctgcattaatgaatcggccaacgcgcggggagag3720gcggtttgcgtattgggcgccagggtggtttttcttttcaccagtgagacgggcaacagc3780tgattgcccttcaccgcctggccctgagagagttgcagcaagcggtccacgctggtttgc3840cccagcaggcgaaaatcctgtttgatggtggttaacggcgggatataacatgagctgtct3900tcggtatcgtcgtatcccactaccgagatatccgcaccaacgcgcagcccggactcggta3960atggcgcgcattgcgcccagcgccatctgatcgttggcaaccagcatcgcagtgggaacg4020atgccctcattcagcatttgcatggtttgttgaaaaccggacatggcactccagtcgcct4080tcccgttccgctatcggctgaatttgattgcgagtgagatatttatgccagccagccaga4140cgcagacgcgccgagacagaacttaatgggcccgctaacagcgcgatttgctggtgaccc4200aatgcgaccagatgctccacgcccagtcgcgtaccgtcttcatgggagaaaataatactg4260ttgatgggtgtctggtcagagacatcaagaaataacgccggaacattagtgcaggcagct4320tccacagcaatggcatcctggtcatccagcggatagttaatgatcagcccactgacgcgt4380tgcgcgagaagattgtgcaccgccgctttacaggcttcgacgccgcttcgttctaccatc4440gacaccaccacgctggcacccagttgatcggcgcgagatttaatcgccgcgacaatttgc4500gacggcgcgtgcagggccagactggaggtggcaacgccaatcagcaacgactgtttgccc4560gccagttgttgtgccacgcggttgggaatgtaattcagctccgccatcgccgcttccact4620ttttcccgcgttttcgcagaaacgtggctggcctggttcaccacgcgggaaacggtctga4680taagagacaccggcatactctgcgacatcgtataacgttactggtttcacattcaccacc4740ctgaattgactctcttccgggcgctatcatgccataccgcgaaaggttttgcgccattcg4800atggtgtccgggatctcgacgctctcccttatgcgactcctgcattaggaagcagcccag4860tagtaggttgaggccgttgagcaccgccgccgcaaggaatggtgcatgcaaggagatggc4920gcccaacagtcccccggccacggggcctgccaccatacccacgccgaaacaagcgctcat4980gagcccgaagtggcgagcccgatcttccccatcggtgatgtcggcgatataggcgccagc5040aaccgcacctgtggcgccggtgatgccggccacgatgcgtccggcgtagaggatcgagat5100ctcgatcccgcgaaattaatacgactcactataggggaattgtgagcggataacaattcc5160cctctagaaataattttgtttaactttaagaaggagatatacatatgacctatactgttg5220gcatgtatcttgcagaacgccttgtacagatcgggctgaagcatcacttcgccgtggcgg5280gcgactacaatctcgttcttctggatcagttgctcctcaacaaggacatgaaacagatct5340attgctgcaatgagttgaactgtggcttcagcgcggaaggctacgcccgttctaacgggg5400ctgcggcagcggttgtcaccttcagcgttggcgccatttccgccatgaacgccctcggcg5460gcgcctatgccgaaaacctgccggttatcctgatttccggcgcgcccaacagcaatgatc5520agggcacaggtcatatcctgcatcacacaatcggcaagacggattacagctaccagcttg5580aaatggcccgtcaggtcacctgtgccgccgaaagcattaccgacgctcactccgccccgg5640ccaagattgaccacgtcattcgcacggcgctgcgcgagcgtaagccggcctatctggaca5700tcgcgtgcaacattgcctccgagccctgcgtgcggcctggccctgtcagcagcctgctgt5760ccgagcctgaaatcgaccacacgagcctgaaggccgcagtggacgccacggttgccttgc5820tggaaaaatcggccagccccgtcatgctgctgggcagcaagctgcgggccgccaacgcac5880tggccgcaaccgaaacgctggcagacaagctgcaatgcgcggtgaccatcatggcggccg5940cgaaaggctttttccccgaagaccacgcgggtttccgcggcctgtactggggcgaagtct6000cgaaccccggcgtgcaggaactggtggagacctccgacgcactgctgtgcatcgcccccg6060tattcaacgactattcaacagtcggctggtcggcatggcccaagggccccaatgtgattc6120tggctgagcccgaccgcgtaacggtcgatggccgcgcctatgacggctttaccctgcgcg6180ccttcctgcaggctctggcggaaaaagcccccgcgcgcccggcctccgcacagaaaagca6240gcgtcccgacgtgctcgctcaccgcgacatccgatgaagccggtctgacgaatgacgaaa6300tcgtccgtcatatcaacgccctgctgacatcaaacacgacgctggtggcagaaaccggcg6360attcatggttcaatgccatgcgcatgaccctgccgcgcggtgcgcgcgtggaactggaaa6420tgcagtggggccatatcggctggtccgtgccctccgccttcggcaatgccatgggctcgc6480aggaccgccagcatgtggtgatggtaggcgatggctccttccagcttaccgcgcaggaag6540tggctcagatggtgcgctacgaactgcccgtcattatctttctgatcaacaaccgtggct6600atgtcggtggcatcgccattcatgacggcccgtacaactatatcaagaactgggattacg6660ccggcctgatggaagtcttcaacgccggagaaggccatggacttggcctgaaagccacca6720ccccgaaggaactgacagaagccatcgccagggcaaaagccaatacccgcggcccgacgc6780tgatcgaatgccagatcgaccgcacggactgcacggatatgctggttcaatggggccgca6840aggttgcctcaaccaacgcgcgcaagaccactctggccctcgagcaccaccaccaccacc6900actgagatccggctgctaacaaagcccgaaaggaagctgagttggctgctgccaccgctg6960agcaataactagcataaccccttggggcctctaaacgggtcttgaggggttttttgctga7020aaggaggaactatatccggat7041<210>6<211>29<212>DNA<213>人工<220><223>人工制备<220><221>misc_feature<222>(14)..(14)<223>n是a、c、g或t<400>6ccgtggctatgtcndtggcatcgccattc29<210>7<211>29<212>DNA<213>人工<220><223>人工制备<220><221>misc_feature<222>(16)..(16)<223>n是a、c、g或t<400>7gaatggcgatgccahngacatagccacgg29<210>8<211>1680<212>DNA<213>人工<220><223>人工制备<400>8atgacctatactgttggcatgtatcttgcagaacgccttgtacagatcgggctgaagcat60cacttcgccgtggcgggcgactacaatctcgttcttctggatcagttgctcctcaacaag120gacatgaaacagatctattgctgcaatgagttgaactgtggcttcagcgcggaaggctac180gcccgttctaacggggctgcggcagcggttgtcaccttcagcgttggcgccatttccgcc240atgaacgccctcggcggcgcctatgccgaaaacctgccggttatcctgatttccggcgcg300cccaacagcaatgatcagggcacaggtcatatcctgcatcacacaatcggcaagacggat360tacagctaccagcttgaaatggcccgtcaggtcacctgtgccgccgaaagcattaccgac420gctcactccgccccggccaagattgaccacgtcattcgcacggcgctgcgcgagcgtaag480ccggcctatctggacatcgcgtgcaacattgcctccgagccctgcgtgcggcctggccct540gtcagcagcctgctgtccgagcctgaaatcgaccacacgagcctgaaggccgcagtggac600gccacggttgccttgctggaaaaatcggccagccccgtcatgctgctgggcagcaagctg660cgggccgccaacgcactggccgcaaccgaaacgctggcagacaagctgcaatgcgcggtg720accatcatggcggccgcgaaaggctttttccccgaagaccacgcgggtttccgcggcctg780tactggggcgaagtctcgaaccccggcgtgcaggaactggtggagacctccgacgcactg840ctgtgcatcgcccccgtattcaacgactattcaacagtcggctggtcggcatggcccaag900ggccccaatgtgattctggctgagcccgaccgcgtaacggtcgatggccgcgcctatgac960ggctttaccctgcgcgccttcctgcaggctctggcggaaaaagcccccgcgcgcccggcc1020tccgcacagaaaagcagcgtcccgacgtgctcgctcaccgcgacatccgatgaagccggt1080ctgacgaatgacgaaatcgtccgtcatatcaacgccctgctgacatcaaacacgacgctg1140gtggcagaaaccggcgattcatggttcaatgccatgcgcatgaccctgccgcgcggtgcg1200cgcgtggaactggaaatgcagtggggccatatcggctggtccgtgccctccgccttcggc1260aatgccatgggctcgcaggaccgccagcatgtggtgatggtaggcgatggctccttccag1320cttaccgcgcaggaagtggctcagatggtgcgctacgaactgcccgtcattatctttctg1380atcaacaaccgtggctatgtcattggcatcgccattcatgacggcccgtacaactatatc1440aagaactgggattacgccggcctgatggaagtcttcaacgccggagaaggccatggactt1500ggcctgaaagccaccaccccgaaggaactgacagaagccatcgccagggcaaaagccaat1560acccgcggcccgacgctgatcgaatgccagatcgaccgcacggactgcacggatatgctg1620gttcaatggggccgcaaggttgcctcaaccaacgcgcgcaagaccactctggccctcgag1680<210>9<211>560<212>PRT<213>人工<220><223>人工制备<400>9MetThrTyrThrValGlyMetTyrLeuAlaGluArgLeuValGlnIle151015GlyLeuLysHisHisPheAlaValAlaGlyAspTyrAsnLeuValLeu202530LeuAspGlnLeuLeuLeuAsnLysAspMetLysGlnIleTyrCysCys354045AsnGluLeuAsnCysGlyPheSerAlaGluGlyTyrAlaArgSerAsn505560GlyAlaAlaAlaAlaValValThrPheSerValGlyAlaIleSerAla65707580MetAsnAlaLeuGlyGlyAlaTyrAlaGluAsnLeuProValIleLeu859095IleSerGlyAlaProAsnSerAsnAspGlnGlyThrGlyHisIleLeu100105110HisHisThrIleGlyLysThrAspTyrSerTyrGlnLeuGluMetAla115120125ArgGlnValThrCysAlaAlaGluSerIleThrAspAlaHisSerAla130135140ProAlaLysIleAspHisValIleArgThrAlaLeuArgGluArgLys145150155160ProAlaTyrLeuAspIleAlaCysAsnIleAlaSerGluProCysVal165170175ArgProGlyProValSerSerLeuLeuSerGluProGluIleAspHis180185190ThrSerLeuLysAlaAlaValAspAlaThrValAlaLeuLeuGluLys195200205SerAlaSerProValMetLeuLeuGlySerLysLeuArgAlaAlaAsn210215220AlaLeuAlaAlaThrGluThrLeuAlaAspLysLeuGlnCysAlaVal225230235240ThrIleMetAlaAlaAlaLysGlyPhePheProGluAspHisAlaGly245250255PheArgGlyLeuTyrTrpGlyGluValSerAsnProGlyValGlnGlu260265270LeuValGluThrSerAspAlaLeuLeuCysIleAlaProValPheAsn275280285AspTyrSerThrValGlyTrpSerAlaTrpProLysGlyProAsnVal290295300IleLeuAlaGluProAspArgValThrValAspGlyArgAlaTyrAsp305310315320GlyPheThrLeuArgAlaPheLeuGlnAlaLeuAlaGluLysAlaPro325330335AlaArgProAlaSerAlaGlnLysSerSerValProThrCysSerLeu340345350ThrAlaThrSerAspGluAlaGlyLeuThrAsnAspGluIleValArg355360365HisIleAsnAlaLeuLeuThrSerAsnThrThrLeuValAlaGluThr370375380GlyAspSerTrpPheAsnAlaMetArgMetThrLeuProArgGlyAla385390395400ArgValGluLeuGluMetGlnTrpGlyHisIleGlyTrpSerValPro405410415SerAlaPheGlyAsnAlaMetGlySerGlnAspArgGlnHisValVal420425430MetValGlyAspGlySerPheGlnLeuThrAlaGlnGluValAlaGln435440445MetValArgTyrGluLeuProValIleIlePheLeuIleAsnAsnArg450455460GlyTyrValGlyGlyIleAlaIleHisAspGlyProTyrAsnTyrIle465470475480LysAsnTrpAspTyrAlaGlyLeuMetGluValPheAsnAlaGlyGlu485490495GlyHisGlyLeuGlyLeuLysAlaThrThrProLysGluLeuThrGlu500505510AlaIleAlaArgAlaLysAlaAsnThrArgGlyProThrLeuIleGlu515520525CysGlnIleAspArgThrAspCysThrAspMetLeuValGlnHisGly530535540ArgLysValAlaSerThrAsnAlaArgLysThrThrLeuAlaLeuGlu545550555560<210>10<211>560<212>PRT<213>人工<220><223>人工制备<400>10MetThrTyrThrValGlyMetTyrLeuAlaGluArgLeuValGlnIle151015GlyLeuLysHisHisPheAlaValAlaGlyAspTyrAsnLeuValLeu202530LeuAspGlnLeuLeuLeuAsnLysAspMetLysGlnIleTyrCysCys354045AsnGluLeuAsnCysGlyPheSerAlaGluGlyTyrAlaArgSerAsn505560GlyAlaAlaAlaAlaValValThrPheSerValGlyAlaIleSerAla65707580MetAsnAlaLeuGlyGlyAlaTyrAlaGluAsnLeuProValIleLeu859095IleSerGlyAlaProAsnSerAsnAspGlnGlyThrGlyHisIleLeu100105110HisHisThrIleGlyLysThrAspTyrSerTyrGlnLeuGluMetAla115120125ArgGlnValThrCysAlaAlaGluSerIleThrAspAlaHisSerAla130135140ProAlaLysIleAspHisValIleArgThrAlaLeuArgGluArgLys145150155160ProAlaTyrLeuAspIleAlaCysAsnIleAlaSerGluProCysVal165170175ArgProGlyProValSerSerLeuLeuSerGluProGluIleAspHis180185190ThrSerLeuLysAlaAlaValAspAlaThrValAlaLeuLeuGluLys195200205SerAlaSerProValMetLeuLeuGlySerLysLeuArgAlaAlaAsn210215220AlaLeuAlaAlaThrGluThrLeuAlaAspLysLeuGlnCysAlaVal225230235240ThrIleMetAlaAlaAlaLysGlyPhePheProGluAspHisAlaGly245250255PheArgGlyLeuTyrTrpGlyGluValSerAsnProGlyValGlnGlu260265270LeuValGluThrSerAspAlaLeuLeuCysIleAlaProValPheAsn275280285AspTyrSerThrValGlyTrpSerAlaTrpProLysGlyProAsnVal290295300IleLeuAlaGluProAspArgValThrValAspGlyArgAlaTyrAsp305310315320GlyPheThrLeuArgAlaPheLeuGlnAlaLeuAlaGluLysAlaPro325330335AlaArgProAlaSerAlaGlnLysSerSerValProThrCysSerLeu340345350ThrAlaThrSerAspGluAlaGlyLeuThrAsnAspGluIleValArg355360365HisIleAsnAlaLeuLeuThrSerAsnThrThrLeuValAlaGluThr370375380GlyAspSerTrpPheAsnAlaMetArgMetThrLeuProArgGlyAla385390395400ArgValGluLeuGluMetGlnTrpGlyHisIleGlyTrpSerValPro405410415SerAlaPheGlyAsnAlaMetGlySerGlnAspArgGlnHisValVal420425430MetValGlyAspGlySerPheGlnLeuThrAlaGlnGluValAlaGln435440445MetValArgTyrGluLeuProValIleIlePheLeuIleAsnAsnArg450455460GlyTyrValGlyGlyIleAlaIleHisAspGlyProTyrAsnTyrIle465470475480LysAsnTrpAspTyrAlaGlyLeuMetGluValPheAsnAlaGlyGlu485490495GlyHisGlyLeuGlyLeuLysAlaThrThrProLysGluLeuThrGlu500505510AlaIleAlaArgAlaLysAlaAsnThrArgGlyProThrLeuIleGlu515520525CysGlnIleAspArgThrAspCysThrAspMetLeuValGlnPheGly530535540ArgLysValAlaSerThrAsnAlaArgLysThrThrLeuAlaLeuGlu545550555560<210>11<211>560<212>PRT<213>人工<220><223>人工制备<400>11MetThrTyrThrValGlyMetTyrLeuAlaGluArgLeuValGlnIle151015GlyLeuLysHisHisPheAlaValAlaGlyAspTyrAsnLeuValLeu202530LeuAspGlnLeuLeuLeuAsnLysAspMetLysGlnIleTyrCysCys354045AsnGluLeuAsnCysGlyPheSerAlaGluGlyTyrAlaArgSerAsn505560GlyAlaAlaAlaAlaValValThrPheSerValGlyAlaIleSerAla65707580MetAsnAlaLeuGlyGlyAlaTyrAlaGluAsnLeuProValIleLeu859095IleSerGlyAlaProAsnSerAsnAspGlnGlyThrGlyHisIleLeu100105110HisHisThrIleGlyLysThrAspTyrSerTyrGlnLeuGluMetAla115120125ArgGlnValThrCysAlaAlaGluSerIleThrAspAlaHisSerAla130135140ProAlaLysIleAspHisValIleArgThrAlaLeuArgGluArgLys145150155160ProAlaTyrLeuAspIleAlaCysAsnIleAlaSerGluProCysVal165170175ArgProGlyProValSerSerLeuLeuSerGluProGluIleAspHis180185190ThrSerLeuLysAlaAlaValAspAlaThrValAlaLeuLeuGluLys195200205SerAlaSerProValMetLeuLeuGlySerLysLeuArgAlaAlaAsn210215220AlaLeuAlaAlaThrGluThrLeuAlaAspLysLeuGlnCysAlaVal225230235240ThrIleMetAlaAlaAlaLysGlyPhePheProGluAspHisAlaGly245250255PheArgGlyLeuTyrTrpGlyGluValSerAsnProGlyValGlnGlu260265270LeuValGluThrSerAspAlaLeuLeuCysIleAlaProValPheAsn275280285AspTyrSerThrValGlyTrpSerAlaTrpProLysGlyProAsnVal290295300IleLeuAlaGluProAspArgValThrValAspGlyArgAlaTyrAsp305310315320GlyPheThrLeuArgAlaPheLeuGlnAlaLeuAlaGluLysAlaPro325330335AlaArgProAlaSerAlaGlnLysSerSerValProThrCysSerLeu340345350ThrAlaThrSerAspGluAlaGlyLeuThrAsnAspGluIleValArg355360365HisIleAsnAlaLeuLeuThrSerAsnThrThrLeuValAlaGluThr370375380GlyAspSerTrpPheAsnAlaMetArgMetThrLeuProArgGlyAla385390395400ArgValGluLeuGluMetGlnTrpGlyHisIleGlyTrpSerValPro405410415SerAlaPheGlyAsnAlaMetGlySerGlnAspArgGlnHisValVal420425430MetValGlyAspGlySerPheGlnLeuThrAlaGlnGluValAlaGln435440445MetValArgTyrGluLeuProValIleIlePheLeuIleAsnAsnArg450455460GlyTyrValGlyGlyIleAlaIleHisAspGlyProTyrAsnTyrIle465470475480LysAsnTrpAspTyrAlaGlyLeuMetGluValPheAsnAlaGlyGlu485490495GlyHisGlyLeuGlyLeuLysAlaThrThrProLysGluLeuThrGlu500505510AlaIleAlaArgAlaLysAlaAsnThrArgGlyProThrLeuIleGlu515520525CysGlnIleAspArgThrAspCysThrAspMetLeuValGlnProGly530535540ArgLysValAlaSerThrAsnAlaArgLysThrThrLeuAlaLeuGlu545550555560<210>12<211>560<212>PRT<213>人工<220><223>人工制备<400>12MetThrTyrThrValGlyMetTyrLeuAlaGluArgLeuValGlnIle151015GlyLeuLysHisHisPheAlaValAlaGlyAspTyrAsnLeuValLeu202530LeuAspGlnLeuLeuLeuAsnLysAspMetLysGlnIleTyrCysCys354045AsnGluLeuAsnCysGlyPheSerAlaGluGlyTyrAlaArgSerAsn505560GlyAlaAlaAlaAlaValValThrPheSerValGlyAlaIleSerAla65707580MetAsnAlaLeuGlyGlyAlaTyrAlaGluAsnLeuProValIleLeu859095IleSerGlyAlaProAsnSerAsnAspGlnGlyThrGlyHisIleLeu100105110HisHisThrIleGlyLysThrAspTyrSerTyrGlnLeuGluMetAla115120125ArgGlnValThrCysAlaAlaGluSerIleThrAspAlaHisSerAla130135140ProAlaLysIleAspHisValIleArgThrAlaLeuArgGluArgLys145150155160ProAlaTyrLeuAspIleAlaCysAsnIleAlaSerGluProCysVal165170175ArgProGlyProValSerSerLeuLeuSerGluProGluIleAspHis180185190ThrSerLeuLysAlaAlaValAspAlaThrValAlaLeuLeuGluLys195200205SerAlaSerProValMetLeuLeuGlySerLysLeuArgAlaAlaAsn210215220AlaLeuAlaAlaThrGluThrLeuAlaAspLysLeuGlnCysAlaVal225230235240ThrIleMetAlaAlaAlaLysGlyPhePheProGluAspHisAlaGly245250255PheArgGlyLeuTyrTrpGlyGluValSerAsnProGlyValGlnGlu260265270LeuValGluThrSerAspAlaLeuLeuCysIleAlaProValPheAsn275280285AspTyrSerThrValGlyTrpSerAlaTrpProLysGlyProAsnVal290295300IleLeuAlaGluProAspArgValThrValAspGlyArgAlaTyrAsp305310315320GlyPheThrLeuArgAlaPheLeuGlnAlaLeuAlaGluLysAlaPro325330335AlaArgProAlaSerAlaGlnLysSerSerValProThrCysSerLeu340345350ThrAlaThrSerAspGluAlaGlyLeuThrAsnAspGluIleValArg355360365HisIleAsnAlaLeuLeuThrSerAsnThrThrLeuValAlaGluThr370375380GlyAspSerTrpPheAsnAlaMetArgMetThrLeuProArgGlyAla385390395400ArgValGluLeuGluMetGlnTrpGlyHisIleGlyTrpSerValPro405410415SerAlaPheGlyAsnAlaMetGlySerGlnAspArgGlnHisValVal420425430MetValGlyAspGlySerPheGlnLeuThrAlaGlnGluValAlaGln435440445MetValArgTyrGluLeuProValIleIlePheLeuIleAsnAsnArg450455460GlyTyrValGlyGlyIleAlaIleHisAspGlyProTyrAsnTyrIle465470475480LysAsnTrpAspTyrAlaGlyLeuMetGluValPheAsnAlaGlyGlu485490495GlyHisGlyLeuGlyLeuLysAlaThrThrProLysGluLeuThrGlu500505510AlaIleAlaArgAlaLysAlaAsnThrArgGlyProThrLeuIleGlu515520525CysGlnIleAspArgThrAspCysThrAspMetLeuValGlnIleGly530535540ArgLysValAlaSerThrAsnAlaArgLysThrThrLeuAlaLeuGlu545550555560<210>13<211>560<212>PRT<213>人工<220><223>人工制备<400>13MetThrTyrThrValGlyMetTyrLeuAlaGluArgLeuValGlnIle151015GlyLeuLysHisHisPheAlaValAlaGlyAspTyrAsnLeuValLeu202530LeuAspGlnLeuLeuLeuAsnLysAspMetLysGlnIleTyrCysCys354045AsnGluLeuAsnCysGlyPheSerAlaGluGlyTyrAlaArgSerAsn505560GlyAlaAlaAlaAlaValValThrPheSerValGlyAlaIleSerAla65707580MetAsnAlaLeuGlyGlyAlaTyrAlaGluAsnLeuProValIleLeu859095IleSerGlyAlaProAsnSerAsnAspGlnGlyThrGlyHisIleLeu100105110HisHisThrIleGlyLysThrAspTyrSerTyrGlnLeuGluMetAla115120125ArgGlnValThrCysAlaAlaGluSerIleThrAspAlaHisSerAla130135140ProAlaLysIleAspHisValIleArgThrAlaLeuArgGluArgLys145150155160ProAlaTyrLeuAspIleAlaCysAsnIleAlaSerGluProCysVal165170175ArgProGlyProValSerSerLeuLeuSerGluProGluIleAspHis180185190ThrSerLeuLysAlaAlaValAspAlaThrValAlaLeuLeuGluLys195200205SerAlaSerProValMetLeuLeuGlySerLysLeuArgAlaAlaAsn210215220AlaLeuAlaAlaThrGluThrLeuAlaAspLysLeuGlnCysAlaVal225230235240ThrIleMetAlaAlaAlaLysGlyPhePheProGluAspHisAlaGly245250255PheArgGlyLeuTyrTrpGlyGluValSerAsnProGlyValGlnGlu260265270LeuValGluThrSerAspAlaLeuLeuCysIleAlaProValPheAsn275280285AspTyrSerThrValGlyTrpSerAlaTrpProLysGlyProAsnVal290295300IleLeuAlaGluProAspArgValThrValAspGlyArgAlaTyrAsp305310315320GlyPheThrLeuArgAlaPheLeuGlnAlaLeuAlaGluLysAlaPro325330335AlaArgProAlaSerAlaGlnLysSerSerValProThrCysSerLeu340345350ThrAlaThrSerAspGluAlaGlyLeuThrAsnAspGluIleValArg355360365HisIleAsnAlaLeuLeuThrSerAsnThrThrLeuValAlaGluThr370375380GlyAspSerTrpPheAsnAlaMetArgMetThrLeuProArgGlyAla385390395400ArgValGluLeuGluMetGlnTrpGlyHisIleGlyTrpSerValPro405410415SerAlaPheGlyAsnAlaMetGlySerGlnAspArgGlnHisValVal420425430MetValGlyAspGlySerPheGlnLeuThrAlaGlnGluValAlaGln435440445MetValArgTyrGluLeuProValIleIlePheLeuIleAsnAsnArg450455460GlyTyrValGlyGlyIleAlaIleHisAspGlyProTyrAsnTyrIle465470475480LysAsnTrpAspTyrAlaGlyLeuMetGluValPheAsnAlaGlyGlu485490495GlyHisGlyLeuGlyLeuLysAlaThrThrProLysGluLeuThrGlu500505510AlaIleAlaArgAlaLysAlaAsnThrArgGlyProThrLeuIleGlu515520525CysGlnIleAspArgThrAspCysThrAspMetLeuValGlnLeuGly530535540ArgLysValAlaSerThrAsnAlaArgLysThrThrLeuAlaLeuGlu545550555560<210>14<211>560<212>PRT<213>人工<220><223>人工制备<400>14MetThrTyrThrValGlyMetTyrLeuAlaGluArgLeuValGlnIle151015GlyLeuLysHisHisPheAlaValAlaGlyAspTyrAsnLeuValLeu202530LeuAspGlnLeuLeuLeuAsnLysAspMetLysGlnIleTyrCysCys354045AsnGluLeuAsnCysGlyPheSerAlaGluGlyTyrAlaArgSerAsn505560GlyAlaAlaAlaAlaValValThrPheSerValGlyAlaIleSerAla65707580MetAsnAlaLeuGlyGlyAlaTyrAlaGluAsnLeuProValIleLeu859095IleSerGlyAlaProAsnSerAsnAspGlnGlyThrGlyHisIleLeu100105110HisHisThrIleGlyLysThrAspTyrSerTyrGlnLeuGluMetAla115120125ArgGlnValThrCysAlaAlaGluSerIleThrAspAlaHisSerAla130135140ProAlaLysIleAspHisValIleArgThrAlaLeuArgGluArgLys145150155160ProAlaTyrLeuAspIleAlaCysAsnIleAlaSerGluProCysVal165170175ArgProGlyProValSerSerLeuLeuSerGluProGluIleAspHis180185190ThrSerLeuLysAlaAlaValAspAlaThrValAlaLeuLeuGluLys195200205SerAlaSerProValMetLeuLeuGlySerLysLeuArgAlaAlaAsn210215220AlaLeuAlaAlaThrGluThrLeuAlaAspLysLeuGlnCysAlaVal225230235240ThrIleMetAlaAlaAlaLysGlyPhePheProGluAspHisAlaGly245250255PheArgGlyLeuTyrTrpGlyGluValSerAsnProGlyValGlnGlu260265270LeuValGluThrSerAspAlaLeuLeuCysIleAlaProValPheAsn275280285AspTyrSerThrValGlyTrpSerAlaTrpProLysGlyProAsnVal290295300IleLeuAlaGluProAspArgValThrValAspGlyArgAlaTyrAsp305310315320GlyPheThrLeuArgAlaPheLeuGlnAlaLeuAlaGluLysAlaPro325330335AlaArgProAlaSerAlaGlnLysSerSerValProThrCysSerLeu340345350ThrAlaThrSerAspGluAlaGlyLeuThrAsnAspGluIleValArg355360365HisIleAsnAlaLeuLeuThrSerAsnThrThrLeuValAlaGluThr370375380GlyAspSerTrpPheAsnAlaMetArgMetThrLeuProArgGlyAla385390395400ArgValGluLeuGluMetGlnTrpGlyHisIleGlyTrpSerValPro405410415SerAlaPheGlyAsnAlaMetGlySerGlnAspArgGlnHisValVal420425430MetValGlyAspGlySerPheGlnLeuThrAlaGlnGluValAlaGln435440445MetValArgTyrGluLeuProValIleIlePheLeuIle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uGlyTyrAlaArgSerAsn505560GlyAlaAlaAlaAlaValValThrPheSerValGlyAlaIleSerAla65707580MetAsnAlaLeuGlyGlyAlaTyrAlaGluAsnLeuProValIleLeu859095IleSerGlyAlaProAsnSerAsnAspGlnGlyThrGlyHisIleLeu100105110HisHisThrIleGlyLysThrAspTyrSerTyrGlnLeuGluMetAla115120125ArgGlnValThrCysAlaAlaGluSerIleThrAspAlaHisSerAla130135140ProAlaLysIleAspHisValIleArgThrAlaLeuArgGluArgLys145150155160ProAlaTyrLeuAspIleAlaCysAsnIleAlaSerGluProCysVal165170175ArgProGlyProValSerSerLeuLeuSerGluProGluIleAspHis180185190ThrSerLeuLysAlaAlaValAspAlaThrValAlaLeuLeuGluLys195200205SerAlaSerProValMetLeuLeuGlySerLysLeuArgAlaAlaAsn210215220AlaLeuAlaAlaThrGluThrLeuAlaAspLysLeuGlnCysAlaVal225230235240ThrIleMetAlaAlaAlaLysGlyPhePheProGluAspHisAlaGly245250255PheArgGlyLeuTyrTrpGlyGluValSerAsnProGlyValGlnGlu260265270LeuValGluThrSerAspAlaLeuLeuCysIleAlaProValPheAsn275280285AspTyrSerThrValGlyTrpSerAlaTrpProLysGlyProAsnVal290295300IleLeuAlaGluProAspArgValThrValAspGlyArgAlaTyrAsp305310315320GlyPheThrLeuArgAlaPheLeuGlnAlaLeuAlaGluLysAlaPro325330335AlaArgProAlaSerAlaGlnLysSerSerValProThrCysSerLeu340345350ThrAlaThrSerAspGluAlaGlyLeuThrAsnAspGluIleValArg355360365HisIleAsnAlaLeuLeuThrSerAsnThrThrLeuValAlaGluThr370375380GlyAspSerTrpPheAsnAlaMetArgMetThrLeuProArgGlyAla385390395400ArgValGluLeuGluMetGlnTrpGlyHisIleGlyTrpSerValPro405410415SerAlaPheGlyAsnAlaMetGlySerGlnAspArgGlnHisValVal420425430MetValGlyAspGlySerPheGlnLeuThrAlaGlnGluValAlaGln435440445MetValArgTyrGluLeuProValIleIlePheLeuIleAsnAsnArg450455460GlyTyrValValGlyIleAlaIleHisAspGlyProTyrAsnTyrIle465470475480LysAsnTrpAspTyrAlaGlyLeuMetGluValPheAsnAlaGlyGlu485490495GlyHisGlyLeuGlyLeuLysAlaThrThrProLysGluLeuThrGlu500505510AlaIleAlaArgAlaLysAlaAsnThrArgGlyProThrLeuIleGlu515520525CysGlnIleAspArgThrAspCysThrAspMetLeuValGlnThrGly530535540ArgLysValAlaSerThrAsnAlaArgLysThrThrLeuAlaLeuGlu545550555560<210>45<211>560<212>PRT<213>人工<220><223>人工制备<400>45MetThrTyrThrValGlyMetTyrLeuAlaGluArgLeuValGlnIle151015GlyLeuLysHisHisPheAlaValAlaGlyAspTyrAsnLeuValLeu202530LeuAspGlnLeuLeuLeuAsnLysAspMetLysGlnIleTyrCysCys354045AsnGluLeuAsnCysGlyPheSerAlaGluGlyTyrAlaArgSerAsn505560GlyAlaAlaAlaAlaValValThrPheSerValGlyAlaIleSerAla65707580MetAsnAlaLeuGlyGlyAlaTyrAlaGluAsnLeuProValIleLeu859095IleSerGlyAlaProAsnSerAsnAspGlnGlyThrGlyHisIleLeu100105110HisHisThrIleGlyLysThrAspTyrSerTyrGlnLeuGluMetAla115120125ArgGlnValThrCysAlaAlaGluSerIleThrAspAlaHisSerAla130135140ProAlaLysIleAspHisValIleArgThrAlaLeuArgGluArgLys145150155160ProAlaTyrLeuAspIleAlaCysAsnIleAlaSerGluProCysVal165170175ArgProGlyProValSerSerLeuLeuSerGluProGluIleAspHis180185190ThrSerLeuLysAlaAlaValAspAlaThrValAlaLeuLeuGluLys195200205SerAlaSerProValMetLeuLeuGlySerLysLeuArgAlaAlaAsn210215220AlaLeuAlaAlaThrGluThrLeuAlaAspLysLeuGlnCysAlaVal225230235240ThrIleMetAlaAlaAlaLysGlyPhePheProGluAspHisAlaGly245250255PheArgGlyLeuTyrTrpGlyGluValSerAsnProGlyValGlnGlu260265270LeuValGluThrSerAspAlaLeuLeuCysIleAlaProValPheAsn275280285AspTyrSerThrValGlyTrpSerAlaTrpProLysGlyProAsnVal290295300IleLeuAlaGluProAspArgValThrValAspGlyArgAlaTyrAsp305310315320GlyPheThrLeuArgAlaPheLeuGlnAlaLeuAlaGluLysAlaPro325330335AlaArgProAlaSerAlaGlnLysSerSerValProThrCysSerLeu340345350ThrAlaThrSerAspGluAlaGlyLeuThrAsnAspGluIleValArg355360365HisIleAsnAlaLeuLeuThrSerAsnThrThrLeuValAlaGluThr370375380GlyAspSerTrpPheAsnAlaMetArgMetThrLeuProArgGlyAla385390395400ArgValGluLeuGluMetGlnTrpGlyHisIleGlyTrpSerValPro405410415SerAlaPheGlyAsnAlaMetGlySerGlnAspArgGlnHisValVal420425430MetValGlyAspGlySerPheGlnLeuThrAlaGlnGluValAlaGln435440445MetValArgTyrGluLeuProValIleIlePheLeuIleAsnAsnArg450455460GlyTyrValValGlyIleAlaIleHisAspGlyProTyrAsnTyrIle465470475480LysAsnTrpAspTyrAlaGlyLeuMetGluValPheAsnAlaGlyGlu485490495GlyHisGlyLeuGlyLeuLysAlaThrThrProLysGluLeuThrGlu500505510AlaIleAlaArgAlaLysAlaAsnThrArgGlyProThrLeuIleGlu515520525CysGlnIleAspArgThrAspCysThrAspMetLeuValGlnGlyGly530535540ArgLysValAlaSerThrAsnAlaArgLysThrThrLeuAlaLeuGlu545550555560<210>46<211>560<212>PRT<213>人工<220><223>人工制备<400>46MetThrTyrThrValGlyMetTyrLeuAlaGluArgLeuValGlnIle151015GlyLeuLysHisHisPheAlaValAlaGlyAspTyrAsnLeuValLeu202530LeuAspGlnLeuLeuLeuAsnLysAspMetLysGlnIleTyrCysCys354045AsnGluLeuAsnCysGlyPheSerAlaGluGlyTyrAlaArgSerAsn505560GlyAlaAlaAlaAlaValValThrPheSerValGlyAlaIleSerAla65707580MetAsnAlaLeuGlyGlyAlaTyrAlaGluAsnLeuProValIleLeu859095IleSerGlyAlaProAsnSerAsnAspGlnGlyThrGlyHisIleLeu100105110HisHisThrIleGlyLysThrAspTyrSerTyrGlnLeuGluMetAla115120125ArgGlnValThrCysAlaAlaGluSerIleThrAspAlaHisSerAla130135140ProAlaLysIleAspHisValIleArgThrAlaLeuArgGluArgLys145150155160ProAlaTyrLeuAspIleAlaCysAsnIleAlaSerGluProCysVal165170175ArgProGlyProValSerSerLeuLeuSerGluProGluIleAspHis180185190ThrSerLeuLysAlaAlaValAspAlaThrValAlaLeuLeuGluLys195200205SerAlaSerProValMetLeuLeuGlySerLysLeuArgAlaAlaAsn210215220AlaLeuAlaAlaThrGluThrLeuAlaAspLysLeuGlnCysAlaVal225230235240ThrIleMetAlaAlaAlaLysGlyPhePheProGluAspHisAlaGly245250255PheArgGlyLeuTyrTrpGlyGluValSerAsnProGlyValGlnGlu260265270LeuValGluThrSerAspAlaLeuLeuCysIleAlaProValPheAsn275280285AspTyrSerThrValGlyTrpSerAlaTrpProLysGlyProAsnVal290295300IleLeuAlaGluProAspArgValThrValAspGlyArgAlaTyrAsp305310315320GlyPheThrLeuArgAlaPheLeuGlnAlaLeuAlaGluLysAlaPro325330335AlaArgProAlaSerAlaGlnLysSerSerValProThrCysSerLeu340345350ThrAlaThrSerAspGluAlaGlyLeuThrAsnAspGluIleValArg355360365HisIleAsnAlaLeuLeuThrSerAsnThrThrLeuValAlaGluThr370375380GlyAspSerTrpPheAsnAlaMetArgMetThrLeuProArgGlyAla385390395400ArgValGluLeuGluMetGlnTrpGlyHisIleGlyTrpSerValPro405410415SerAlaPheGlyAsnAlaMetGlySerGlnAspArgGlnHisValVal420425430MetValGlyAspGlySerPheGlnLeuThrAlaGlnGluValAlaGln435440445MetValArgTyrGluLeuProValIleIlePheLeuIleAsnAsnArg450455460GlyTyrValValGlyIleAlaIleHisAspGlyProTyrAsnTyrIle465470475480LysAsnTrpAspTyrAlaGlyLeuMetGluValPheAsnAlaGlyGlu485490495GlyHisGlyLeuGlyLeuLysAlaThrThrProLysGluLeuThrGlu500505510AlaIleAlaArgAlaLysAlaAsnThrArgGlyProThrLeuIleGlu515520525CysGlnIleAspArgThrAspCysThrAspMetLeuValGlnSerGly530535540ArgLysValAlaSerThrAsnAlaArgLysThrThrLeuAlaLeuGlu545550555560<210>47<211>560<212>PRT<213>人工<220><223>人工制备<400>47MetThrTyrThrValGlyMetTyrLeuAlaGluArgLeuValGlnIle151015GlyLeuLysHisHisPheAlaValAlaGlyAspTyrAsnLeuValLeu202530LeuAspGlnLeuLeuLeuAsnLysAspMetLysGlnIleTyrCysCys354045AsnGluLeuAsnCysGlyPheSerAlaGluGlyTyrAlaArgSerAsn505560GlyAlaAlaAlaAlaValValThrPheSerValGlyAlaIleSerAla65707580MetAsnAlaLeuGlyGlyAlaTyrAlaGluAsnLeuProValIleLeu859095IleSerGlyAlaProAsnSerAsnAspGlnGlyThrGlyHisIleLeu100105110HisHisThrIleGlyLysThrAspTyrSerTyrGlnLeuGluMetAla115120125ArgGlnValThrCysAlaAlaGluSerIleThrAspAlaHisSerAla130135140ProAlaLysIleAspHisValIleArgThrAlaLeuArgGluArgLys145150155160ProAlaTyrLeuAspIleAlaCysAsnIleAlaSerGluProCysVal165170175ArgProGlyProValSerSerLeuLeuSerGluProGluIleAspHis180185190ThrSerLeuLysAlaAlaValAspAlaThrValAlaLeuLeuGluLys195200205SerAlaSerProValMetLeuLeuGlySerLysLeuArgAlaAlaAsn210215220AlaLeuAlaAlaThrGluThrLeuAlaAspLysLeuGlnCysAlaVal225230235240ThrIleMetAlaAlaAlaLysGlyPhePheProGluAspHisAlaGly245250255PheArgGlyLeuTyrTrpGlyGluValSerAsnProGlyValGlnGlu260265270LeuValGluThrSerAspAlaLeuLeuCysIleAlaProValPheAsn275280285AspTyrSerThrValGlyTrpSerAlaTrpProLysGlyProAsnVal290295300IleLeuAlaGluProAspArgValThrValAspGlyArgAlaTyrAsp305310315320GlyPheThrLeuArgAlaPheLeuGlnAlaLeuAlaGluLysAlaPro325330335AlaArgProAlaSerAlaGlnLysSerSerValProThrCysSerLeu340345350ThrAlaThrSerAspGluAlaGlyLeuThrAsnAspGluIleValArg355360365HisIleAsnAlaLeuLeuThrSerAsnThrThrLeuValAlaGluThr370375380GlyAspSerTrpPheAsnAlaMetArgMetThrLeuProArgGlyAla385390395400ArgValGluLeuGluMetGlnTrpGlyHisIleGlyTrpSerValPro405410415SerAlaPheGlyAsnAlaMetGlySerGlnAspArgGlnHisValVal420425430MetValGlyAspGlySerPheGlnLeuThrAlaGlnGluValAlaGln435440445MetValArgTyrGluLeuProValIleIlePheLeuIleAsnAsnArg450455460GlyTyrValValGlyIleAlaIleHisAspGlyProTyrAsnTyrIle465470475480LysAsnTrpAspTyrAlaGlyLeuMetGluValPheAsnAlaGlyGlu485490495GlyHisGlyLeuGlyLeuLysAlaThrThrProLysGluLeuThrGlu500505510AlaIleAlaArgAlaLysAlaAsnThrArgGlyProThrLeuIleGlu515520525CysGlnIleAspArgThrAspCysThrAspMetLeuValGlnCysGly530535540ArgLysValAlaSerThrAsnAlaArgLysThrThrLeuAlaLeuGlu545550555560<210>48<211>1680<212>DNA<213>人工<220><223>人工制备<400>48atgacctatactgttggcatgtatcttgcagaacgccttgtacagatcgggctgaagcat60cacttcgccgtggcgggcgactacaatctcgttcttctggatcagttgctcctcaacaag120gacatgaaacagatctattgctgcaatgagttgaactgtggcttcagcgcggaaggctac180gcccgttctaacggggctgcggcagcggttgtcaccttcagcgttggcgccatttccgcc240atgaacgccctcggcggcgcctatgccgaaaacctgccggttatcctgatttccggcgcg300cccaacagcaatgatcagggcacaggtcatatcctgcatcacacaatcggcaagacggat360tacagctaccagcttgaaatggcccgtcaggtcacctgtgccgccgaaagcattaccgac420gctcactccgccccggccaagattgaccacgtcattcgcacggcgctgcgcgagcgtaag480ccggcctatctggacatcgcgtgcaacattgcctccgagccctgcgtgcggcctggccct540gtcagcagcctgctgtccgagcctgaaatcgaccacacgagcctgaaggccgcagtggac600gccacggttgccttgctggaaaaatcggccagccccgtcatgctgctgggcagcaagctg660cgggccgccaacgcactggccgcaaccgaaacgctggcagacaagctgcaatgcgcggtg720accatcatggcggccgcgaaaggctttttccccgaagaccacgcgggtttccgcggcctg780tactggggcgaagtctcgaaccccggcgtgcaggaactggtggagacctccgacgcactg840ctgtgcatcgcccccgtattcaacgactattcaacagtcggctggtcggcatggcccaag900ggccccaatgtgattctggctgagcccgaccgcgtaacggtcgatggccgcgcctatgac960ggctttaccctgcgcgccttcctgcaggctctggcggaaaaagcccccgcgcgcccggcc1020tccgcacagaaaagcagcgtcccgacgtgctcgctcaccgcgacatccgatgaagccggt1080ctgacgaatgacgaaatcgtccgtcatatcaacgccctgctgacatcaaacacgacgctg1140gtggcagaaaccggcgattcatggttcaatgccatgcgcatgaccctgccgcgcggtgcg1200cgcgtggaactggaaatgcagtggggccatatcggctggtccgtgccctccgccttcggc1260aatgccatgggctcgcaggaccgccagcatgtggtgatggtaggcgatggctccttccag1320cttaccgcgcaggaagtggctcagatggtgcgctacgaactgcccgtcattatctttctg1380atcaacaaccgtggctatgtcgccggcatcgccattcatgacggcccgtacaactatatc1440aagaactgggattacgccggcctgatggaagtcttcaacgccggagaaggccatggactt1500ggcctgaaagccaccaccccgaaggaactgacagaagccatcgccagggcaaaagccaat1560acccgcggcccgacgctgatcgaatgccagatcgaccgcacggactgcacggatatgctg1620gttcaatttggccgcaaggttgcctcaaccaacgcgcgcaagaccactctggccctcgag1680<210>49<211>7041<212>DNA<213>人工<220><223>人工制备<400>49tggcgaatgggacgcgccctgtagcggcgcattaagcgcggcgggtgtggtggttacgcg60cagcgtgaccgctacacttgccagcgccctagcgcccgctcctttcgctttcttcccttc120ctttctcgccacgttcgccggctttccccgtcaagctctaaatcgggggctccctttagg180gttccgatttagtgctttacggcacctcgaccccaaaaaacttgattagggtgatggttc240acgtagtgggccatcgccctgatagacggtttttcgccctttgacgttggagtccacgtt300ctttaatagtggactcttgttccaaactggaacaacactcaaccctatctcggtctattc360ttttgatttataagggattttgccgatttcggcctattggttaaaaaatgagctgattta420acaaaaatttaacgcgaattttaacaaaatattaacgtttacaatttcaggtggcacttt480tcggggaaatgtgcgcggaacccctatttgtttatttttctaaatacattcaaatatgta540tccgctcatgagacaataaccctgataaatgcttcaataatattgaaaaaggaagagtat600gagtattcaacatttccgtgtcgcccttattcccttttttgcggcattttgccttcctgt660ttttgctcacccagaaacgctggtgaaagtaaaagatgctgaagatcagttgggtgcacg720agtgggttacatcgaactggatctcaacagcggtaagatccttgagagttttcgccccga780agaacgttttccaatgatgagcacttttaaagttctgctatgtggcgcggtattatcccg840tattgacgccgggcaagagcaactcggtcgccgcatacactattctcagaatgacttggt900tgagtactcaccagtcacagaaaagcatcttacggatggcatgacagtaagagaattatg960cagtgctgccataaccatgagtgataacactgcggccaacttacttctgacaacgatcgg1020aggaccgaaggagctaaccgcttttttgcacaacatgggggatcatgtaactcgccttga1080tcgttgggaaccggagctgaatgaagccataccaaacgacgagcgtgacaccacgatgcc1140tgcagcaatggcaacaacgttgcgcaaactattaactggcgaactacttactctagcttc1200ccggcaacaattaatagactggatggaggcggataaagttgcaggaccacttctgcgctc1260ggcccttccggctggctggtttattgctgataaatctggagccggtgagcgtgggtctcg1320cggtatcattgcagcactggggccagatggtaagccctcccgtatcgtagttatctacac1380gacggggagtcaggcaactatggatgaacgaaatagacagatcgctgagataggtgcctc1440actgattaagcattggtaactgtcagaccaagtttactcatatatactttagattgattt1500aaaacttcatttttaatttaaaaggatctaggtgaagatcctttttgataatctcatgac1560caaaatcccttaacgtgagttttcgttccactgagcgtcagaccccgtagaaaagatcaa1620aggatcttcttgagatcctttttttctgcgcgtaatctgctgcttgcaaacaaaaaaacc1680accgctaccagcggtggtttgtttgccggatcaagagctaccaactctttttccgaaggt1740aactggcttcagcagagcgcagataccaaatactgtccttctagtgtagccgtagttagg1800ccaccacttcaagaactctgtagcaccgcctacatacctcgctctgctaatcctgttacc1860agtggctgctgccagtggcgataagtcgtgtcttaccgggttggactcaagacgatagtt1920accggataaggcgcagcggtcgggctgaacggggggttcgtgcacacagcccagcttgga1980gcgaacgacctacaccgaactgagatacctacagcgtgagctatgagaaagcgccacgct2040tcccgaagggagaaaggcggacaggtatccggtaagcggcagggtcggaacaggagagcg2100cacgagggagcttccagggggaaacgcctggtatctttatagtcctgtcgggtttcgcca2160cctctgacttgagcgtcgatttttgtgatgctcgtcaggggggcggagcctatggaaaaa2220cgccagcaacgcggcctttttacggttcctggccttttgctggccttttgctcacatgtt2280ctttcctgcgttatcccctgattctgtggataaccgtattaccgcctttgagtgagctga2340taccgctcgccgcagccgaacgaccgagcgcagcgagtcagtgagcgaggaagcggaaga2400gcgcctgatgcggtattttctccttacgcatctgtgcggtatttcacaccgcatatatgg2460tgcactctcagtacaatctgctctgatgccgcatagttaagccagtatacactccgctat2520cgctacgtgactgggtcatggctgcgccccgacacccgccaacacccgctgacgcgccct2580gacgggcttgtctgctcccggcatccgcttacagacaagctgtgaccgtctccgggagct2640gcatgtgtcagaggttttcaccgtcatcaccgaaacgcgcgaggcagctgcggtaaagct2700catcagcgtggtcgtgaagcgattcacagatgtctgcctgttcatccgcgtccagctcgt2760tgagtttctccagaagcgttaatgtctggcttctgataaagcgggccatgttaagggcgg2820ttttttcctgtttggtcactgatgcctccgtgtaagggggatttctgttcatgggggtaa2880tgataccgatgaaacgagagaggatgctcacgatacgggttactgatgatgaacatgccc2940ggttactggaacgttgtgagggtaaacaactggcggtatggatgcggcgggaccagagaa3000aaatcactcagggtcaatgccagcgcttcgttaatacagatgtaggtgttccacagggta3060gccagcagcatcctgcgatgcagatccggaacataatggtgcagggcgctgacttccgcg3120tttccagactttacgaaacacggaaaccgaagaccattcatgttgttgctcaggtcgcag3180acgttttgcagcagcagtcgcttcacgttcgctcgcgtatcggtgattcattctgctaac3240cagtaaggcaaccccgccagcctagccgggtcctcaacgacaggagcacgatcatgcgca3300cccgtggggccgccatgccggcgataatggcctgcttctcgccgaaacgtttggtggcgg3360gaccagtgacgaaggcttgagcgagggcgtgcaagattccgaataccgcaagcgacaggc3420cgatcatcgtcgcgctccagcgaaagcggtcctcgccgaaaatgacccagagcgctgccg3480gcacctgtcctacgagttgcatgataaagaagacagtcataagtgcggcgacgatagtca3540tgccccgcgcccaccggaaggagctgactgggttgaaggctctcaagggcatcggtcgag3600atcccggtgcctaatgagtgagctaacttacattaattgcgttgcgctcactgcccgctt3660tccagtcgggaaacctgtcgtgccagctgcattaatgaatcggccaacgcgcggggagag3720gcggtttgcgtattgggcgccagggtggtttttcttttcaccagtgagacgggcaacagc3780tgattgcccttcaccgcctggccctgagagagttgcagcaagcggtccacgctggtttgc3840cccagcaggcgaaaatcctgtttgatggtggttaacggcgggatataacatgagctgtct3900tcggtatcgtcgtatcccactaccgagatatccgcaccaacgcgcagcccggactcggta3960atggcgcgcattgcgcccagcgccatctgatcgttggcaaccagcatcgcagtgggaacg4020atgccctcattcagcatttgcatggtttgttgaaaaccggacatggcactccagtcgcct4080tcccgttccgctatcggctgaatttgattgcgagtgagatatttatgccagccagccaga4140cgcagacgcgccgagacagaacttaatgggcccgctaacagcgcgatttgctggtgaccc4200aatgcgaccagatgctccacgcccagtcgcgtaccgtcttcatgggagaaaataatactg4260ttgatgggtgtctggtcagagacatcaagaaataacgccggaacattagtgcaggcagct4320tccacagcaatggcatcctggtcatccagcggatagttaatgatcagcccactgacgcgt4380tgcgcgagaagattgtgcaccgccgctttacaggcttcgacgccgcttcgttctaccatc4440gacaccaccacgctggcacccagttgatcggcgcgagatttaatcgccgcgacaatttgc4500gacggcgcgtgcagggccagactggaggtggcaacgccaatcagcaacgactgtttgccc4560gccagttgttgtgccacgcggttgggaatgtaattcagctccgccatcgccgcttccact4620ttttcccgcgttttcgcagaaacgtggctggcctggttcaccacgcgggaaacggtctga4680taagagacaccggcatactctgcgacatcgtataacgttactggtttcacattcaccacc4740ctgaattgactctcttccgggcgctatcatgccataccgcgaaaggttttgcgccattcg4800atggtgtccgggatctcgacgctctcccttatgcgactcctgcattaggaagcagcccag4860tagtaggttgaggccgttgagcaccgccgccgcaaggaatggtgcatgcaaggagatggc4920gcccaacagtcccccggccacggggcctgccaccatacccacgccgaaacaagcgctcat4980gagcccgaagtggcgagcccgatcttccccatcggtgatgtcggcgatataggcgccagc5040aaccgcacctgtggcgccggtgatgccggccacgatgcgtccggcgtagaggatcgagat5100ctcgatcccgcgaaattaatacgactcactataggggaattgtgagcggataacaattcc5160cctctagaaataattttgtttaactttaagaaggagatatacatatgacctatactgttg5220gcatgtatcttgcagaacgccttgtacagatcgggctgaagcatcacttcgccgtggcgg5280gcgactacaatctcgttcttctggatcagttgctcctcaacaaggacatgaaacagatct5340attgctgcaatgagttgaactgtggcttcagcgcggaaggctacgcccgttctaacgggg5400ctgcggcagcggttgtcaccttcagcgttggcgccatttccgccatgaacgccctcggcg5460gcgcctatgccgaaaacctgccggttatcctgatttccggcgcgcccaacagcaatgatc5520agggcacaggtcatatcctgcatcacacaatcggcaagacggattacagctaccagcttg5580aaatggcccgtcaggtcacctgtgccgccgaaagcattaccgacgctcactccgccccgg5640ccaagattgaccacgtcattcgcacggcgctgcgcgagcgtaagccggcctatctggaca5700tcgcgtgcaacattgcctccgagccctgcgtgcggcctggccctgtcagcagcctgctgt5760ccgagcctgaaatcgaccacacgagcctgaaggccgcagtggacgccacggttgccttgc5820tggaaaaatcggccagccccgtcatgctgctgggcagcaagctgcgggccgccaacgcac5880tggccgcaaccgaaacgctggcagacaagctgcaatgcgcggtgaccatcatggcggccg5940cgaaaggctttttccccgaagaccacgcgggtttccgcggcctgtactggggcgaagtct6000cgaaccccggcgtgcaggaactggtggagacctccgacgcactgctgtgcatcgcccccg6060tattcaacgactattcaacagtcggctggtcggcatggcccaagggccccaatgtgattc6120tggctgagcccgaccgcgtaacggtcgatggccgcgcctatgacggctttaccctgcgcg6180ccttcctgcaggctctggcggaaaaagcccccgcgcgcccggcctccgcacagaaaagca6240gcgtcccgacgtgctcgctcaccgcgacatccgatgaagccggtctgacgaatgacgaaa6300tcgtccgtcatatcaacgccctgctgacatcaaacacgacgctggtggcagaaaccggcg6360attcatggttcaatgccatgcgcatgaccctgccgcgcggtgcgcgcgtggaactggaaa6420tgcagtggggccatatcggctggtccgtgccctccgccttcggcaatgccatgggctcgc6480aggaccgccagcatgtggtgatggtaggcgatggctccttccagcttaccgcgcaggaag6540tggctcagatggtgcgctacgaactgcccgtcattatctttctgatcaacaaccgtggct6600atgtcgccggcatcgccattcatgacggcccgtacaactatatcaagaactgggattacg6660ccggcctgatggaagtcttcaacgccggagaaggccatggacttggcctgaaagccacca6720ccccgaaggaactgacagaagccatcgccagggcaaaagccaatacccgcggcccgacgc6780tgatcgaatgccagatcgaccgcacggactgcacggatatgctggttcaatttggccgca6840aggttgcctcaaccaacgcgcgcaagaccactctggccctcgagcaccaccaccaccacc6900actgagatccggctgctaacaaagcccgaaaggaagctgagttggctgctgccaccgctg6960agcaataactagcataaccccttggggcctctaaacgggtcttgaggggttttttgctga7020aaggaggaactatatccggat7041<210>50<211>34<212>DNA<213>人工<220><223>人工制备<400>50accttgcggccgaattgaaccagcatatccgtgc34<210>51<211>34<212>DNA<213>人工<220><223>人工制备<400>51gcacggatatgctggttcaattcggccgcaaggt34<210>52<211>1680<212>DNA<213>人工<220><223>人工制备<400>52atgacctatactgttggcatgtatcttgcagaacgccttgtacagatcgggctgaagcat60cacttcgccgtggcgggcgactacaatctcgttcttctggatcagttgctcctcaacaag120gacatgaaacagatctattgctgcaatgagttgaactgtggcttcagcgcggaaggctac180gcccgttctaacggggctgcggcagcggttgtcaccttcagcgttggcgccatttccgcc240atgaacgccctcggcggcgcctatgccgaaaacctgccggttatcctgatttccggcgcg300cccaacagcaatgatcagggcacaggtcatatcctgcatcacacaatcggcaagacggat360tacagctaccagcttgaaatggcccgtcaggtcacctgtgccgccgaaagcattaccgac420gctcactccgccccggccaagattgaccacgtcattcgcacggcgctgcgcgagcgtaag480ccggcctatctggacatcgcgtgcaacattgcctccgagccctgcgtgcggcctggccct540gtcagcagcctgctgtccgagcctgaaatcgaccacacgagcctgaaggccgcagtggac600gccacggttgccttgctggaaaaatcggccagccccgtcatgctgctgggcagcaagctg660cgggccgccaacgcactggccgcaaccgaaacgctggcagacaagctgcaatgcgcggtg720accatcatggcggccgcgaaaggctttttccccgaagaccacgcgggtttccgcggcctg780tactggggcgaagtctcgaaccccggcgtgcaggaactggtggagacctccgacgcactg840ctgtgcatcgcccccgtattcaacgactattcaacagtcggctggtcggcatggcccaag900ggccccaatgtgattctggctgagcccgaccgcgtaacggtcgatggccgcgcctatgac960ggctttaccctgcgcgccttcctgcaggctctggcggaaaaagcccccgcgcgcccggcc1020tccgcacagaaaagcagcgtcccgacgtgctcgctcaccgcgacatccgatgaagccggt1080ctgacgaatgacgaaatcgtccgtcatatcaacgccctgctgacatcaaacacgacgctg1140gtggcagaaaccggcgattcatggttcaatgccatgcgcatgaccctgccgcgcggtgcg1200cgcgtggaactggaaatgcagtggggccatatcggctggtccgtgccctccgccttcggc1260aatgccatgggctcgcaggaccgccagcatgtggtgatggtaggcgatggctccttccag1320cttaccgcgcaggaagtggctcagatggtgcgctacgaactgcccgtcattatctttctg1380atcaacaaccgtggctatgtcgccggcatcgccattcatgacggcccgtacaactatatc1440aagaactgggattacgccggcctgatggaagtcttcaacgccggagaaggccatggactt1500ggcctgaaagccaccaccccgaaggaactgacagaagccatcgccagggcaaaagccaat1560acccgcggcccgacgctgatcgaatgccagatcgaccgcacggactgcacggatatgctg1620gttcaatggggccgcaaggttgcctcaaccaacgcgcgcaagaccactctggccctcgag1680<210>53<211>560<212>PRT<213>人工<220><223>人工制备<400>53MetThrTyrThrValGlyMetTyrLeuAlaGluArgLeuValGlnIle151015GlyLeuLysHisHisPheAlaValAlaGlyAspTyrAsnLeuValLeu202530LeuAspGlnLeuLeuLeuAsnLysAspMetLysGlnIleTyrCysCys354045AsnGluLeuAsnCysGlyPheSerAlaGluGlyTyrAlaArgSerAsn505560GlyAlaAlaAlaAlaValValThrPheSerValGlyAlaIleSerAla65707580MetAsnAlaLeuGlyGlyAlaTyrAlaGluAsnLeuProValIleLeu859095IleSerGlyAlaProAsnSerAsnAspGlnGlyThrGlyHisIleLeu100105110HisHisThrIleGlyLysThrAspTyrSerTyrGlnLeuGluMetAla115120125ArgGlnValThrCysAlaAlaGluSerIleThrAspAlaHisSerAla130135140ProAlaLysIleAspHisValIleArgThrAlaLeuArgGluArgLys145150155160ProAlaTyrLeuAspIleAlaCysAsnIleAlaSerGluProCysVal165170175ArgProGlyProValSerSerLeuLeuSerGluProGluIleAspHis180185190ThrSerLeuLysAlaAlaValAspAlaThrValAlaLeuLeuGluLys195200205SerAlaSerProValMetLeuLeuGlySerLysLeuArgAlaAlaAsn210215220AlaLeuAlaAlaThrGluThrLeuAlaAspLysLeuGlnCysAlaVal225230235240ThrIleMetAlaAlaAlaLysGlyPhePheProGluAspHisAlaGly245250255PheArgGlyLeuTyrTrpGlyGluValSerAsnProGlyValGlnGlu260265270LeuValGluThrSerAspAlaLeuLeuCysIleAlaProValPheAsn275280285AspTyrSerThrValGlyTrpSerAlaTrpProLysGlyProAsnVal290295300IleLeuAlaGluProAspArgValThrValAspGlyArgAlaTyrAsp305310315320GlyPheThrLeuArgAlaPheLeuGlnAlaLeuAlaGluLysAlaPro325330335AlaArgProAlaSerAlaGlnLysSerSerValProThrCysSerLeu340345350ThrAlaThrSerAspGluAlaGlyLeuThrAsnAspGluIleValArg355360365HisIleAsnAlaLeuLeuThrSerAsnThrThrLeuValAlaGluThr370375380GlyAspSerTrpPheAsnAlaMetArgMetThrLeuProArgGlyAla385390395400ArgValGluLeuGluMetGlnTrpGlyHisIleGlyTrpSerValPro405410415SerAlaPheGlyAsnAlaMetGlySerGlnAspArgGlnHisValVal420425430MetValGlyAspGlySerPheGlnLeuThrAlaGlnGluValAlaGln435440445MetValArgTyrGluLeuProValIleIlePheLeuIleAsnAsnArg450455460GlyTyrValIleGlyIleAlaIleHisAspGlyProTyrAsnTyrIle465470475480LysAsnTrpAspTyrAlaGlyLeuMetGluValPheAsnAlaGlyGlu485490495GlyHisGlyLeuGlyLeuLysAlaThrThrProLysGluLeuThrGlu500505510AlaIleAlaArgAlaLysAlaAsnThrArgGlyProThrLeuIleGlu515520525CysGlnIleAspArgThrAspCysThrAspMetLeuValGlnTrpGly530535540ArgLysValAlaSerThrAsnAlaArgLysThrThrLeuAlaLeuGlu545550555560当前第1页1 2 3