專利名稱：1－脫氧－d－木酮糖生物合成途徑的基因的制作方法
技術(shù)領(lǐng)域：
本發(fā)明涉及DNA序列，當(dāng)將該DNA摻入到病毒、真核生物和原核生物基因組時，所述的DNA序列修飾類異戊二烯的生物合成，并且涉及生產(chǎn)這些轉(zhuǎn)基因病毒、真核生物和原核生物的遺傳工程方法。本發(fā)明還涉及鑒定在植物中具有除草、抗微生物、抗寄生蟲、抗病毒、殺真菌、殺細(xì)菌的作用和在人體和動物中具有抗微生物、抗寄生蟲、抗真菌、抗細(xì)菌和抗病毒的作用的物質(zhì)的方法。
已經(jīng)知道借助于經(jīng)典的乙酸/甲羥戊酸途徑或者甲羥戊酸-依賴性生物合成途徑，脫氧-D-木酮糖磷酸途徑形成類異戊二烯的生物合成途徑(Rohmer,M.,Knani,M.,Simonin,P.,Sutter,B.和Sahm,H.(1993)生物化學(xué)雜志295:517-524)。
但是，借助于脫氧-D-木酮糖磷酸途徑如何和通過怎樣的途徑可能引起病毒、真核生物和原核生物中類異戊二烯濃度的變化是未知的。

圖1顯示該生物合成途徑。
因此提供了DNA序列，該DNA序列編碼1-脫氧-D-木酮糖5-磷酸合酶(DOXP合酶)、1-脫氧-D-木酮糖-5-磷酸還原異構(gòu)酶(DOXP還原異構(gòu)酶)或gcpE蛋白。所有三個基因和酶都與類異戊二烯生物合成有關(guān)。
gcpE蛋白具有激酶功能并催化糖或磷糖類或類異戊二烯生物合成的前體的磷酸化，特別是2-C-甲基-D-赤蘚糖醇、2-C-甲基-D-赤蘚糖醇磷酸酯的磷酸化，特別是2-C-甲基-D-赤蘚糖醇-4-磷酸酯、2-C-甲基-D-赤蘚糖、2-C-甲基-D-赤蘚糖磷酸酯的磷酸化，特別是2-C-甲基-D-赤蘚糖-4-磷酸酯的磷酸化。對于類異戊二烯合成的前體，gcpE蛋白特別地催化下面物質(zhì)的磷酸化CH2(OH)-C(CH3)=C(OH)-CH2-O-PO(OH)2,CH2(OH)-C(CH3)=C(OH)-CH2-OH,CH2(OH)-CH(CH3)-CO-CH2-O-PO(OH)2,CH2(OH)-CH(CH3)-CO-CH2OH,CH2=C(CH3)-CO-CH2-O-PO(OH)2,CH2=C(CH3)-CO-CH2-OH,CH2=C(CH3)-CH(OH)-CH2-O-PO(OH)2,CH2=C(CH3)-CH(OH)-CH2-OH,CH2(OH)-C(=CH2)-C(OH)-CH2-O-PO(OH)2,CH2(OH)-C(=CH2)-C(OH)-CH2-OH,CHO-CH(CH3)-CH(OH)-CH2-O-PO-(OH)2,CHO-CH(CH3)-CH(OH)-CH2-OH,CH2(OH)-C(OH)(CH3)-CH=CH-O-PO(OH)2,CH2(OH)-C(OH)(CH3)-CH=CH-OHCH(OH)=C(CH3)-CH(OH)-CH2-O-PO(OH)2,CH(OH)=C(CH3)-CH(OH)-CH2-OH(CH3)2HC-CO-CH2-O-PO(OH)2,(CH3)2HC-CO-CH2-O-H,(CH3)2HC-CH(OH)-CH2-O-PO(OH)2,(CH3)2HC-CH(OH)-CH2-O-H。
DOXP合酶催化丙酮酸和甘油醛-3-磷酸縮合產(chǎn)生1-脫氧-D-木酮糖-5-磷酸酯和DOXP還原異構(gòu)酶催化1-脫氧-D-木酮糖-5-磷酸酯轉(zhuǎn)變?yōu)?-C-甲基-D-赤蘚糖醇-4-磷酸酯(參見圖1)。
本發(fā)明涉及下面的DNA序列編碼具有SEQ ID No.2顯示的氨基酸序列的多肽或編碼SEQ ID No.2的多肽的類似物或者衍生物的DNA序列，該類似物或衍生物是其中一個或多個氨基酸缺失、添加或被其它氨基酸替代；編碼具有SEQ ID No.4顯示的氨基酸序列的多肽或編碼SEQ ID No.4的多肽的類似物或者衍生物的DNA序列，該類似物或衍生物是其中一個或多個氨基酸缺失、添加或被其它氨基酸替代；編碼具有SEQ ID No.6顯示的氨基酸序列的多肽或編碼SEQ ID No.6的多肽的類似物或者衍生物的DNA序列，該類似物或衍生物是其中一個或多個氨基酸缺失、添加或被其它氨基酸替代。
該基因和其基因產(chǎn)物(多肽)以它們的一級結(jié)構(gòu)表示并被指定如下SEQ ID No.1:1-脫氧-D-木酮糖-5-磷酸還原異構(gòu)酶基因SEQ ID No.2:1-脫氧-D-木酮糖-5-磷酸還原異構(gòu)酶SEQ ID No.3:1-脫氧-D-木酮糖-5-磷酸合酶基因SEQ ID No.4:1-脫氧-D-木酮糖-5-磷酸合酶SEQ ID No.5:gcpE基因SEQ ID No.6:gcpE蛋白。
所有DNA序列都來源于惡性瘧原蟲。
除了在序列表中列出的DNA序列，合適的序列也包括作為遺傳密碼簡并的結(jié)果的具有不同DNA序列但是編碼相同的肽或編碼該多肽的類似物或者衍生物的序列，該衍生物是其中一個或多個氨基酸序列缺失、添加或被其它氨基酸替代。
本發(fā)明的序列適用于在病毒、真核生物和原核生物中基因的表達(dá)，該基因負(fù)責(zé)通過1-脫氧-D-木酮糖途徑的類異戊二烯的生物合成。
根據(jù)本發(fā)明，真核生物或者真核細(xì)胞包括動物細(xì)胞、植物細(xì)胞、藻類、酵母、真菌，而原核生物或原核細(xì)胞包括細(xì)菌、古細(xì)菌和真細(xì)菌。
當(dāng)將DNA序列摻入到定位了上面所述DNA序列的基因組時，上面所說的基因能夠在病毒、真核生物和原核生物中表達(dá)。以本領(lǐng)域內(nèi)技術(shù)人員已知的方式培養(yǎng)根據(jù)本發(fā)明轉(zhuǎn)化的病毒、真核生物和原核生物并且分離和任選地純化在這樣的培養(yǎng)期間形成的類異戊二烯。不是所有的類異戊二烯都需要分離，因為在一些情況下，類異戊二烯被直接釋放到環(huán)境空氣中。
此外，本發(fā)明涉及生產(chǎn)轉(zhuǎn)基因病毒、真核生物和原核生物以改變類異戊二烯含量的方法，該方法包括下面步驟a)生產(chǎn)具有下面亞序列的DNA序列ⅰ)在病毒、真核生物和原核生物中具有活性并且在預(yù)定的靶組織或靶細(xì)胞中確保形成RNA的啟動子，ⅱ)編碼具有顯示于SEQ ID No.2、4或6的氨基酸序列的多肽或者SEQ ID No.2、4或6的多肽的類似物或者衍生物的DNA序列，ⅲ)能夠或者加強(qiáng)所述基因在病毒、真核生物和原核生物中表達(dá)的5’和3’未翻譯的序列，b)利用或不利用載體(例如質(zhì)粒、病毒DNA)將該DNA序列轉(zhuǎn)移和摻入到病毒、原核或真核細(xì)胞的基因組中。
完整的植物可以從這種方式轉(zhuǎn)化的植物細(xì)胞再生。
可以使具有SEQ ID No.1、SEQ ID No.3和SEQ ID No.5的核苷酸序列的蛋白質(zhì)編碼序列含有確保在某些器官或細(xì)胞中轉(zhuǎn)錄的啟動子，該啟動子以有義方向偶合(啟動子的3’末端到編碼序列的5’末端)到編碼待形成的蛋白質(zhì)的序列。將決定mRNA合成的終止信號連接到編碼序列的3’末端。為了將待表達(dá)的蛋白質(zhì)導(dǎo)入某些亞細(xì)胞區(qū)室，例如葉綠體、造粉體、線粒體、液泡、細(xì)胞溶膠或者細(xì)胞間隙，可以將編碼所謂的信號序列或轉(zhuǎn)移肽的其它的序列插入到啟動子和編碼序列之間。在一些情況下，插入編碼蛋白質(zhì)的羧基末端信號的序列是必要的。該序列必須是在與該蛋白質(zhì)的編碼序列相同的閱讀框架中。為了準(zhǔn)備將本發(fā)明的DNA序列導(dǎo)入到高等植物中需要獲得大量的克隆載體，所述的載體含有大腸桿菌的復(fù)制信號和用以選擇轉(zhuǎn)化細(xì)胞的標(biāo)記物。根據(jù)將需要的基因?qū)氲街参镏械姆椒?，可能需要其它的DNA序列。例如如果將Ti和Ri質(zhì)粒用于轉(zhuǎn)化植物細(xì)胞，必須插入Ti和Ri質(zhì)粒T-DNA的至少一個右邊界，但是通常是右邊界和左邊界，作為待導(dǎo)入基因的側(cè)面區(qū)域。在歐洲專利120516；Hoekama在“二性植物載體系統(tǒng)”，Offset-drukkerij KantersB.V.Alblasserdam(1985)，第5章；Fraley等，植物科學(xué)的關(guān)鍵和綜述4,1-46和An等(1985)EMBO J.4,277-287中廣泛地調(diào)查和全面地描述了T-DNA在轉(zhuǎn)化植物細(xì)胞中的用途。一旦將導(dǎo)入的DNA摻入到基因組中，它通常是穩(wěn)定的并且還保留在原始轉(zhuǎn)化細(xì)胞的子代中。通常它含有選擇性標(biāo)記，該標(biāo)記給予轉(zhuǎn)化植物細(xì)胞對殺生物劑或抗生素，例如卡那霉素、G418、博來霉素、潮霉素或磷蘇菌素等的抗性。因而所使用的特別的標(biāo)記物傾向于允許從缺乏插入的DNA的細(xì)胞中選擇出轉(zhuǎn)化細(xì)胞。
許多技術(shù)可用于將DNA導(dǎo)入到植物。這些技術(shù)包括用農(nóng)桿菌例如根癌農(nóng)桿菌的輔助轉(zhuǎn)化、原生質(zhì)體融合、DNA的微注射、電穿孔，以及微彈轟擊方法和病毒注射。然后整個植物可以在合適的培養(yǎng)基中從轉(zhuǎn)化植物材料再生，所述的培養(yǎng)基可以含有用于選擇目的的抗生素或殺生物劑。對于用于注射和電穿孔的質(zhì)粒沒有特別的要求。但是，如果完整植株要從這樣的轉(zhuǎn)化細(xì)胞再生，必須存在選擇性標(biāo)記物基因。轉(zhuǎn)化細(xì)胞以常規(guī)的方式在植物中生長(McCormick等(1986)，植物細(xì)胞報告5,81-84)?？梢哉Ｅ囵B(yǎng)該植物并且與具有相同的轉(zhuǎn)化基因組或其它基因組的植物進(jìn)行雜交。獲得的個體具有相應(yīng)的表現(xiàn)型特性。
本發(fā)明還提供了表達(dá)載體，它含有一個或多個本發(fā)明的DNA序列。通過使本發(fā)明的DNA序列含有合適的功能調(diào)節(jié)信號獲得這樣的表達(dá)載體。這樣的調(diào)節(jié)信號是負(fù)責(zé)表達(dá)的DNA序列，例如啟動子、操縱基因、增強(qiáng)子、核糖體結(jié)合位點(diǎn)，并且被宿主生物體識別。
其它的調(diào)節(jié)信號例如控制重組DNA在宿主生物體中復(fù)制或重組的調(diào)節(jié)信號，也可以任選地是表達(dá)載體的組成部份。
本發(fā)明還提供了用本發(fā)明的DNA序列或表達(dá)載體轉(zhuǎn)化的宿主生物體。
用于表達(dá)本發(fā)明的酶的合適的宿主細(xì)胞和生物體是那些不包括具有DOXP合酶、DOXP還原異構(gòu)酶或gcpE蛋白功能的固有酶的生物體。這是對古細(xì)菌、動物、真菌、粘菌和一些真細(xì)菌的情況。這樣的固有酶活性的缺失本質(zhì)上簡化了重組酶的檢測和純化。因此，也是首次可能通過各種化學(xué)品和藥物直接在宿主細(xì)胞的粗提物中測定本發(fā)明的重組酶的活性和特別是活性的抑制。
如果想要實(shí)現(xiàn)多肽鏈的翻譯后修飾和天然折疊，那么本發(fā)明的酶有利地在真核細(xì)胞中表達(dá)。此外，取決于表達(dá)系統(tǒng)，當(dāng)表達(dá)基因組序列時，確保通過剪接DNA去除內(nèi)含子，并產(chǎn)生對寄生蟲獨(dú)特的的多肽序列的酶。利用重組DNA技術(shù)，編碼內(nèi)含子的序列可以被從待表達(dá)的DNA序列去除或為了試驗的目的被插入到待表達(dá)的DNA序列中。
利用本領(lǐng)域技術(shù)人員已知的方法可以從宿主細(xì)胞或宿主細(xì)胞的培養(yǎng)物上清液分離蛋白質(zhì)。也可能需要酶的體外再激活。
為了簡化純化，可以以具有不同肽鏈的融合蛋白表達(dá)本發(fā)明的酶或該酶的亞序列。寡聚-組氨酸序列和來源于谷胱甘肽S-轉(zhuǎn)移酶、硫氧還蛋白或鈣調(diào)蛋白結(jié)合肽的序列是特別適用于該目的。
此外，可以以具有本領(lǐng)域技術(shù)人員已知的這樣的肽鏈的融合蛋白來表達(dá)本發(fā)明的酶或該酶的亞序列，該重組酶被運(yùn)輸?shù)桨馀囵B(yǎng)基或運(yùn)輸?shù)剿拗骷?xì)胞的某些區(qū)室。因此簡化了該酶的純化和生物學(xué)活性的研究。
當(dāng)表達(dá)本發(fā)明的酶時，可以證明修飾個體密碼子是方便的。如果在寄生蟲中使用的密碼子不同于在異源表達(dá)系統(tǒng)中使用的密碼子，該編碼區(qū)域中有目的的堿基的替代也是可取的，以確保蛋白質(zhì)的最佳合成。
此外，可以在標(biāo)準(zhǔn)化的條件下，采用本領(lǐng)域技術(shù)人員已知的方法進(jìn)行體外翻譯獲得本發(fā)明的酶。適用于這一目的的系統(tǒng)是兔網(wǎng)織紅細(xì)胞和小麥胚芽提取物和細(xì)菌溶解產(chǎn)物。也可以將體外轉(zhuǎn)錄mRNA翻譯到非洲爪蟾屬卵母細(xì)胞。
其序列來源于本發(fā)明的酶的肽序列的寡肽和多肽可以通過化學(xué)合成獲得。如果合適地選擇序列，這樣的肽具有本發(fā)明酶特征的特性。這樣的肽可以大量地產(chǎn)生并且特別適合于研究酶活性動力學(xué)、酶活性的調(diào)節(jié)、酶的三維結(jié)構(gòu)，各種化學(xué)品和藥物對酶活性的抑制，以及各種配體的結(jié)合幾何學(xué)和結(jié)合親和力。
優(yōu)選地將具有SEQ ID No.1、3和5的序列的核苷酸的DNA用于重組生產(chǎn)本發(fā)明的酶。
因此本發(fā)明還涉及用于篩選抑制脫氧-D-木酮糖磷酸代謝途徑的化合物的方法。根據(jù)該方法，提供了含有重組表達(dá)載體的宿主生物體，其中載體包括至少SEQ ID No.1、SEQ ID No.3或SEQ ID No.5的寡核苷酸序列或其變體或同系物的一部分，和提供了被懷疑對人和動物具有抗微生物、抗寄生蟲、抗細(xì)菌、抗病毒和抗真菌的作用或?qū)χ参锞哂锌刮⑸铩⒖共《?、殺?xì)菌、除草或殺真菌的活性的化合物。然后將宿主生物體與該化合物接觸并測定化合物的活性。
本發(fā)明也提供了用于測定gcpE蛋白的酶促活性的方法。利用已知的方法可以測定所述的活性。通過測定糖或磷糖或類異戊二烯生物合成的前體的磷酸化，特別是2-C-甲基-D-赤蘚糖醇、2-C-甲基-D-赤蘚糖醇磷酸酯的磷酸化，特別是2-C-甲基-D-赤蘚糖醇-4-磷酸酯、2-C-甲基-D-赤蘚糖、2-C-甲基-D-赤蘚糖磷酸酯的磷酸化，特別是2-C-甲基-D-赤蘚糖-4-磷酸至的磷酸化進(jìn)行測定。本發(fā)明還提供了該測定方法在識別抑制特別的酶的活性的物質(zhì)中的用途。
通過測定甘油醛-3-磷酸向2-C-甲基赤蘚糖醇-4-磷酸酯的轉(zhuǎn)變，可以單步測定DOXP合酶和DOXP還原異構(gòu)酶的酶促活性。
類似地進(jìn)行DOXP合酶和DOXP還原異構(gòu)酶的酶促活性的測定。由Querol等描述的熒光法也適用于測定DOXP合酶活性(Querol等，摘要，第4屆歐洲植物類異戊二烯研討會，巴塞羅那，1999年4月21-23日)。
序列表<110>哈桑·朱馬<120>1-脫氧-D-木酮糖生物合成途徑的基因<130>15696<140>PCT/EP99<141>1999-09-22<150>DE19923567.8<151>1999-05-22<150>DE19843279.8<151>1998-09-22<160>6<170>PatentIn Ver.2.1<210>1<211>1467<212>DNA<213>惡性瘧原蟲<220><221>CDS<222>(1)..(1467)<220><221>基因<222>(1)..(1467)<220><221>mRNA<222>(1)..(1467)<400>1atg aag aaa tat att tat ata tat ttt ttc ttc atc aca ata act att 48Met Lys Lys Tyr Ile Tyr Ile Tyr Phe Phe Phe Ile Thr Ile Thr Ile1 5 10 15aat gat tta gta ata aat aat aca tca aaa tgt gtt tcc att gaa aga 96Asn Asp Leu Val Ile Asn Asn Thr Ser Lys Cys Val Ser Ile Glu Arg20 25 30aga aaa aat aac gca tat ata aat tat ggt ata gga tat aat gga cca 144Arg Lys Asn Asn Ala Tyr Ile Asn Tyr Gly Ile Gly Tyr Asn Gly Pro35 40 45gat aat aaa ata aca aag agt aga aga tgt aaa aga ata aag tta tgc 192Asp Asn Lys Ile Thr Lys Ser Arg Arg Cys Lys Arg Ile Lys Leu Cys50 55 60aaa aag gat tta ata gat att ggt gca ata aag aaa cca att aat gta 240Lys Lys Asp Leu Ile Asp Ile Gly Ala Ile Lys Lys Pro Ile Asn Val65 70 75 80gca att ttt gga agt act ggt agt ata ggt acg aat gct tta aat ata 288Ala Ile Phe Gly Ser Thr Gly Ser Ile Gly Thr Asn Ala Leu Asn Ile85 90 95ata agg gag tgt aat aaa att gaa aat gtt ttt aat gtt aaa gca ttg 336Ile Arg Glu Cys Asn Lys Ile Glu Asn Val Phe Asn Val Lys Ala Leu100 105 110tat gtg aat aag agt gtg aat gaa tta tat gaa caa gct aga gaa ttt 384Tyr Val Asn Lys Ser Val Asn Glu Leu Tyr Glu Gln Ala Arg Glu Phe115 120 125tta cca gaa tat ttg tgt ata cat gat aaa agt gta tat gaa gaa tta 432Leu Pro Glu Tyr Leu Cys Ile His Asp Lys Ser Val Tyr Glu Glu Leu130 135 140aaa gaa ctg gta aaa aat ata aaa gat tat aaa cct ata ata ttg tgt 480Lys Glu Leu Val Lys Asn Ile Lys Asp Tyr Lys Pro Ile Ile Leu Cys145 150 155 160ggt gat gaa ggg atg aaa gaa ata tgt agt agt aat agt ata gat aaa 528Gly Asp Glu Gly Met Lys Glu Ile Cys Ser Ser Asn Ser Ile Asp Lys165 170 175ata gtt att ggt att gat tct ttt caa gga tta tat tct act atg tat 576Ile Val Ile Gly Ile Asp Ser Phe Gln Gly Leu Tyr Ser Thr Met Tyr180 185 190gca att atg aat aat aaa ata gtt gcg tta gct aat aaa gaa tcc att 624Ala Ile Met Asn Asn Lys Ile Val Ala Leu Ala Asn Lys Glu Ser Ile195 200 205gtc tct gct ggt ttc ttt tta aag aaa tta tta aat att cat aaa aat 672Val Ser Ala Gly Phe Phe Leu Lys Lys Leu Leu Asn Ile His Lys Asn210 215 220gca aag ata ata cct gtt gat tca gaa cat agt gct ata ttt caa tgt 720Ala Lys Ile Ile Pro Val Asp Ser Glu His Ser Ala Ile Phe Gln Cys225 230 235 240tta gat aat aat aag gta tta aaa aca aaa tgt tta caa gac aat ttt 768Leu Asp Asn Asn Lys Val Leu Lys Thr Lys Cys Leu Gln Asp Asn Phe245 250 255tct aaa att aac aat ata aat aaa ata ttt tta tgt tca tct gga ggt 816Ser Lys Ile Asn Asn Ile Asn Lys Ile Phe Leu Cys Ser Ser Gly Gly260 265 270cca ttt caa aat tta act atg gac gaa tta aaa aat gta aca tca gaa 864Pro Phe Gln Asn Leu Thr Met Asp Glu Leu Lys Asn Val Thr Ser Glu275 280 285aat gct tta aag cat cct aaa tgg aaa atg ggt aag aaa ata act ata 912Asn Ala Leu Lys His Pro Lys Trp Lys Met Gly Lys Lys Ile Thr Ile290 295 300gat tct gca act atg atg aat aaa ggt tta gag gtt ata gaa acc cat 960Asp Ser Ala Thr Met Met Asn Lys Gly Leu Glu Val Ile Glu Thr His305 310 315 320ttt tta ttt gat gta gat tat aat gat ata gaa gtt ata gta cat aaa 1008Phe Leu Phe Asp Val Asp Tyr Asn Asp Ile Glu Val Ile Val His Lys325 330 335gaa tgc att ata cat tct tgt gtt gaa ttt ata gac aaa tca gta ata 1056Glu Cys Ile Ile His Ser Cys Val Glu Phe Ile Asp Lys Ser Val Ile340 345 350agt caa atg tat tat cca gat atg caa ata ccc ata tta tat tct tta 1104Ser Gln Met Tyr Tyr Pro Asp Met Gln Ile Pro Ile Leu Tyr Ser Leu355 360 365aca tgg cct gat aga ata aaa aca aat tta aaa cct tta gat ttg gct 1152Thr Trp Pro Asp Arg Ile Lys Thr Asn Leu Lys Pro Leu Asp Leu Ala370 375 380cag gtt tca act ctt aca ttt cat aaa cct tct tta gaa cat ttc ccg 1200Gln Val Ser Thr Leu Thr Phe His Lys Pro Ser Leu Glu His Phe Pro385 390 395 400tgt att aaa tta gct tat caa gca ggt ata aaa gga aac ttt tat cca 1248Cys Ile Lys Leu Ala Tyr Gln Ala Gly Ile Lys Gly Asn Phe Tyr Pro405410 415act gta cta aat gcg tca aat gaa ata gct aac aac tta ttt ttg aat 1296Thr Val Leu Asn Ala Ser Asn Glu Ile Ala Asn Asn Leu Phe Leu Asn420 425 430aat aaa att aaa tat ttt gat att tcc tct ata ata tcg caa gtt ctt 1344Asn Lys Ile Lys Tyr Phe Asp Ile Ser Ser Ile Ile Ser Gln Val Leu435 440 445gaa tct ttc aat tct caa aag gtt tcg gaa aat agt gaa gat tta atg 1392Glu Ser Phe Asn Ser Gln Lys Val Ser Glu Asn Ser Glu Asp Leu Met450 455 460aag caa att cta caa ata cat tct tgg gcc aaa gat aaa gct acc gat 1440Lys Gln Ile Leu Gln Ile His Ser Trp Ala Lys Asp Lys Ala Thr Asp465 470 475 480ata tac aac aaa cat aat tct tca tag 1467Ile Tyr Asn Lys His Asn Ser Ser485<210>2<211>488<212>PRT<213>惡性瘧原蟲<400>2Met Lys Lys Tyr Ile Tyr Ile Tyr Phe Phe Phe Ile Thr Ile Thr Ile1 5 10 15Asn Asp Leu Val Ile Asn Asn Thr Ser Lys Cys Val Ser Ile Glu Arg20 25 30Arg Lys Asn Asn Ala Tyr Ile Asn Tyr Gly Ile Gly Tyr Asn Gly Pro35 40 45Asp Asn Lys Ile Thr Lys Ser Arg Arg Cys Lys Arg Ile Lys Leu Cys50 55 60Lys Lys Asp Leu Ile Asp Ile Gly Ala Ile Lys Lys Pro Ile Asn Val65 70 75 80Ala Ile Phe Gly Ser Thr Gly Ser Ile Gly Thr Asn Ala Leu Asn Ile85 90 95Ile Arg Glu Cys Asn Lys Ile Glu Asn Val Phe Asn Val Lys Ala Leu100 105 110Tyr Val Asn Lys Ser Val Asn Glu Leu Tyr Glu Gln Ala Arg Glu Phe115 120 125Leu Pro Glu Tyr Leu Cys Ile His Asp Lys Ser Val Tyr Glu Glu Leu130 135 140Lys Glu Leu Val Lys Asn Ile Lys Asp Tyr Lys Pro Ile Ile Leu Cys145 150 155 160Gly Asp Glu Gly Met Lys Glu Ile Cys Ser Ser Asn Ser Ile Asp Lys165 170 175Ile Val Ile Gly Ile Asp Ser Phe Gln Gly Leu Tyr Ser Thr Met Tyr180 185 190Ala Ile Met Asn Asn Lys Ile Val Ala Leu Ala Asn Lys Glu Ser Ile195 200 205Val Ser Ala Gly Phe Phe Leu Lys Lys Leu Leu Asn Ile His Lys Asn210 215 220Ala Lys Ile Ile Pro Val Asp Ser Glu His Ser Ala Ile Phe Gln Cys225 230 235 240Leu Asp Asn Asn Lys Val Leu Lys Thr Lys Cys Leu Gln Asp Asn Phe245 250 255Ser Lys Ile Asn Asn Ile Asn Lys Ile Phe Leu Cys Ser Ser Gly Gly260 265 270Pro Phe Gln Asn Leu Thr Met Asp Glu Leu Lys Asn Val Thr Ser Glu275 280 285Asn Ala Leu Lys His Pro Lys Trp Lys Met Gly Lys Lys Ile Thr Ile290 295 300Asp Ser Ala Thr Met Met Asn Lys Gly Leu Glu Val Ile Glu Thr His305 310 315 320Phe Leu Phe Asp Val Asp Tyr Asn Asp Ile Glu Val Ile Val His Lys325 330 335Glu Cys Ile Ile His Ser Cys Val Glu Phe Ile Asp Lys Ser Val Ile340 345 350Ser Gln Met Tyr Tyr Pro Asp Met Gln Ile Pro Ile Leu Tyr Ser Leu355 360 365Thr Trp Pro Asp Arg Ile Lys Thr Asn Leu Lys Pro Leu Asp Leu Ala370 375 380Gln Val Ser Thr Leu Thr Phe His Lys Pro Ser Leu Glu His Phe Pro385 390 395 400Cys Ile Lys Leu Ala Tyr Gln Ala Gly Ile Lys Gly Asn Phe Tyr Pro405 410 415Thr Val Leu Asn Ala Ser Asn Glu Ile Ala Asn Asn Leu Phe Leu Asn420 425 430Asn Lys Ile Lys Tyr Phe Asp Ile Ser Ser Ile Ile Ser Gln Val Leu435 440 445Glu Ser Phe Asn Ser Gln Lys Val Ser Glu Asn Ser Glu Asp Leu Met450 455 460Lys Gln Ile Leu Gln Ile His Ser Trp Ala Lys Asp Lys Ala Thr Asp465 470 475 480Ile Tyr Asn Lys His Asn Ser Ser485<210>3<211>3872<212>DNA<213>惡性瘧原蟲<220><221>CDS<222>(126)..(3740)<220><221>基因<222>(1)..(3870)<220><221>mRNA<222>(1)..(3870)<400>3ggtaatatac gtataatata tatataatat attcttacgt atgtatcatt tatgaatcat 60aataatattc taaatttacc ttccgttttt gctcgatctt ctcattttcg tttcagcttt 120tatca atg att ttt aat tat gtg ttt ttt aag aac ttt gta cca gtt gtt 170Met Ile Phe Asn Tyr Val Phe Phe Lys Asn Phe Val Pro Val Val1 5 10 15cta tac att ctc ctt ata ata tat att aac tta aat ggc atg aat aat 218Leu Tyr Ile Leu Leu Ile Ile Tyr Ile Asn Leu Asn Gly Met Asn Asn20 25 30aaa aat caa ata aaa aca gaa aaa att tat ata aag aaa ttg aat agg 266Lys Asn Gln Ile Lys Thr Glu Lys Ile Tyr Ile Lys Lys Leu Asn Arg35 40 45ttg tca agg aaa aat tcg tta tgt agt tct aaa aat aaa ata gca tgc 314Leu Ser Arg Lys Asn Ser Leu Cys Ser Ser Lys Asn Lys Ile Ala Cys50 55 60ttg ttc gat ata gga aat gat gat aat aga aat acg aca tat ggc tat 362Leu Phe Asp Ile Gly Asn Asp Asp Asn Arg Asn Thr Thr Tyr Gly Tyr65 70 75aat gtg aat gtt aaa aat gat gat att aat tcc tta cta aaa aat aat 410Asn Val Asn Val Lys Asn Asp Asp Ile Asn Ser Leu Leu Lys Asn Asn80 85 90 95tat agt aat aaa ttg tac atg gat aag agg aaa aat att aat aat gta 458Tyr Ser Asn Lys Leu Tyr Met Asp Lys Arg Lys Asn Ile Asn Asn Val100 105 110att agt act aat aaa ata tct ggg tcc att tca aat att tgt agt aga 506Ile Ser Thr Asn Lys Ile Ser Gly Ser Ile Ser Asn Ile Cys Ser Arg115 120 125aat caa aaa gaa aat gaa caa aaa aga aat aaa caa aga tgt tta act 554Asn Gln Lys Glu Asn Glu Gln Lys Arg Asn Lys Gln Arg Cys Leu Thr130 135 140caa tgt cac act tat aat atg tca cat gaa cag gac aaa cta gct aat 602Gln Cys His Thr Tyr Asn Met Ser His Glu Gln Asp Lys Leu Ala Asn145 150 155gat aat aat agg aat aat aaa aag aat ttt aat tta tta ttt ata aat 650Asp Asn Asn Arg Asn Asn Lys Lys Asn Phe Asn Leu Leu Phe Ile Asn160 165 170 175tat ttt aat ttg aaa cga atg aaa aat tct ctt cta aat aaa gac aat 698Tyr Phe Asn Leu Lys Arg Met Lys Asn Ser Leu Leu Asn Lys Asp Asn180 185 190ttc ttt tac tgt aaa gaa aaa aaa ttg tca ttt ctg cat aag gcc tat 746Phe Phe Tyr Cys Lys Glu Lys Lys Leu Ser Phe Leu His Lys Ala Tyr195 200 205aaa aaa aaa aat tgc act ttt caa aat tat agt tta aaa aga aaa tct 794Lys Lys Lys Asn Cys Thr Phe Gln Asn Tyr Ser Leu Lys Arg Lys Ser210 215 220aat cgt gat tca cat aaa ttg ttt tct gga gaa ttt gac gat tat aca 842Asn Arg Asp Ser His Lys Leu Phe Ser Gly Glu Phe Asp Asp Tyr Thr225 230 235aat aat aat gct tta tat gaa tcc gaa aaa aaa gaa tac att aca cta 890Asn Asn Asn Ala Leu Tyr Glu Ser Glu Lys Lys Glu Tyr Ile Thr Leu240 245 250 255aat aat aat aat aaa aat aat aat aat aaa aat aat gat aat aaa aat 938Asn Asn Asn Asn Lys Asn Asn Asn Asn Lys Asn Asn Asp Asn Lys Asn260 265 270aat gat aat aat gat tat aat aat aat aat agt tgt aat aat tta gga 986Asn Asp Asn Asn Asp Tyr Asn Asn Asn Asn Ser Cys Asn Asn Leu Gly275 280 285gag aga tcc aat cat tat gat aat tat ggt gga gat aat aat aat cca 1034Glu Arg Ser Asn His Tyr Asp Asn Tyr Gly Gly Asp Asn Asn Asn Pro290 295 300tgt aat aat aat aat gac aaa tat gat ata gga aaa tat ttc aaa cag 1082Cys Asn Asn Asn Asn Asp Lys Tyr Asp Ile Gly Lys Tyr Phe Lys Gln305 310 315att aat acc ttt att aat att gat gaa tat aaa act ata tat ggt gat 1130Ile Asn Thr Phe Ile Asn Ile Asp Glu Tyr Lys Thr Ile Tyr Gly Asp320 325 330 335gaa ata tat aaa gaa ata tat gaa cta tat gta gaa aga aat att cct 1178Glu Ile Tyr Lys Glu Ile Tyr Glu Leu Tyr Val Glu Arg Asn Ile Pro340 345 350gaa tat tat gaa cga aaa tat ttt tca gaa gat att aaa aag agt gtc 1226Glu Tyr Tyr Glu Arg Lys Tyr Phe Ser Glu Asp Ile Lys Lys Set Val355 360 365cta ttt gat ata gat aaa tat aat gat gtc gaa ttt gaa aaa gct ata 1274Leu Phe Asp Ile Asp Lys Tyr Asn Asp Val Glu Phe Glu Lys Ala Ile370 375 380aaa gaa gaa ttt ata aat aat gga gtt tat att aat aat ata gat aat 1322Lys Glu Glu Phe Ile Asn Asn Gly Val Tyr Ile Asn Asn Ile Asp Asn385 390 395aca tat tat aaa aaa gaa aat att tta ata atg aaa aag ata tta cat 1370Thr Tyr Tyr Lys Lys Glu Asn Ile Leu Ile Met Lys Lys Ile Leu His400 405 410 415tat ttc cca tta tta aaa tta att aat aat cca tca gat tta aaa aag 1418Tyr Phe Pro Leu Leu Lys Leu Ile Asn Asn Pro Ser Asp Leu Lys Lys420 425 430tta aaa aaa caa tat tta cct tta tta gca cat gaa tta aaa ata ttt 1466Leu Lys Lys Gln Tyr Leu Pro Leu Leu Ala His Glu Leu Lys Ile Phe435 440 445tta ttt ttt att gta aat ata aca gga ggt cat ttt tcc tct gtt tta 1514Leu Phe Phe Ile Val Asn Ile Thr Gly Gly His Phe Ser Ser Val Leu450 455 460agc tct tta gaa att caa tta tta tta ttg tat att ttt aat caa cca 1562Ser Ser Leu Glu Ile Gln Leu Leu Leu Leu Tyr Ile Phe Asn Gln Pro465 470 475tat gat aat gtt ata tat gat ata gga cat caa gca tat gta cat aag 1610Tyr Asp Asn Val Ile Tyr Asp Ile Gly His Gln Ala Tyr Val His Lys480 485 490 495ata ttg acc gga aga aaa cta tta ttt cta tca tta aga aat aaa aaa 1658Ile Leu Thr Gly Arg Lys Leu Leu Phe Leu Ser Leu Arg Asn Lys Lys500 505 510ggt att agt gga ttc cta aat att ttt gaa agt att tat gat aaa ttt 1706Gly Ile Ser Gly Phe Leu Asn Ile Phe Glu Ser Ile Tyr Asp Lys Phe515 520 525ggg gct ggt cac agt tcc act tca tta agt gct ata caa gga tat tat 1754Gly Ala Gly His Ser Ser Thr Ser Leu Ser Ala Ile Gln Gly Tyr Tyr530 535 540gaa gcc gag tgg caa gtg aag aat aaa gaa aaa tat gga aat gga gat 1802Glu Ala Glu Trp Gln Val Lys Asn Lys Glu Lys Tyr Gly Asn Gly Asp545 550 555ata gaa ata agt gat aac gca aat gtc acg aat aat gaa agg ata ttt 1850Ile Glu Ile Ser Asp Asn Ala Asn Val Thr Asn Asn Glu Arg Ile Phe560 565 570 575caa aaa gga ata cac aat gat aat aat att aac aat aat att aat aat 1898Gln Lys Gly Ile His Asn Asp Asn Asn Ile Asn Asn Asn Ile Asn Asn580 585 590aat aat tat atc aat cct tca gat gtg gta gga aga gaa aat acg aat 1946Asn Asn Tyr Ile Asn Pro Ser Asp Val Val Gly Arg Glu Asn Thr Asn595 600 605gta cca aat gta cga aat gat aac cat aac gtg gat aaa gta cac att 1994Val Pro Asn Val Arg Asn Asp Asn His Asn Val Asp Lys Val His Ile610 615 620gct att ata gga gat ggt ggt tta aca ggt gga atg gca tta gaa gcg 2042Ala Ile Ile Gly Asp Gly Gly Leu Thr Gly Gly Met Ala Leu Glu Ala625 630 635tta aat tat att tca ttc ttg aat tct aaa att tta att att tat aat 2090Leu Asn Tyr Ile Ser Phe Leu Asn Ser Lys Ile Leu Ile Ile Tyr Asn640 645 650 655gat aac gga caa gtt tct tta cca aca aat gcc gta agt ata tca ggt 2138Asp Asn Gly Gln Val Ser Leu Pro Thr Asn Ala Val Ser Ile Ser Gly660 665 670aat aga cct ata ggt tct ata tca gat cat tta cat tat ttt gtt tct 2186Asn Arg Pro Ile Gly Ser Ile Ser Asp His Leu His Tyr Phe Val Ser675 680 685aat ata gaa gca aat gct ggt gat aat aaa tta tcg aaa aat gca aaa 2234Asn Ile Glu Ala Asn Ala Gly Asp Asn Lys Leu Ser Lys Asn Ala Lys690 695 700gag aat aac att ttt gaa aat ttg aat tat gat tat att ggt gtt gtg 2282Glu Asn Asn Ile Phe Glu Asn Leu Asn Tyr Asp Tyr Ile Gly Val Val705 710 715aat ggt aat aat aca gaa gag ctc ttt aaa gta tta aat aat ata aaa 2330Asn Gly Asn Asn Thr Glu Glu Leu Phe Lys Val Leu Asn Asn Ile Lys720 725 730 735gaa aat aaa tta aaa aga gct act gtt ctt cat gta cgt aca aaa aaa 2378Glu Asn Lys Leu Lys Arg Ala Thr Val Leu His Val Arg Thr Lys Lys740 745 750tcg aat gat ttt ata aat tca aag agt cca ata agt ata ttg cac tct 2426Ser Asn Asp Phe Ile Asn Ser Lys Ser Pro Ile Ser Ile Leu His Ser755 760 765ata aag aaa aat gag att ttc cct ttc gat acc act ata tta aat gga 2474Ile Lys Lys Asn Glu Ile Phe Pro Phe Asp Thr Thr Ile Leu Asn Gly770 775 780aat att cat aag gag aac aag ata gaa gaa gag aaa aat gtg tct tca 2522Asn Ile His Lys Glu Asn Lys Ile Glu Glu Glu Lys Asn Val Ser Ser785 790 795tct aca aag tat gat gta aat aat aag aat aat aaa aat aat gat aat 2570Ser Thr Lys Tyr Asp Val Asn Asn Lys Asn Asn Lys Asn Asn Asp Asn800 805 810 815agt gaa att ata aaa tat gaa gat atg ttt tca aaa gag acg ttc aca 2618Ser Glu Ile Ile Lys Tyr Glu Asp Met Phe Ser Lys Glu Thr Phe Thr820 825 830gat ata tat aca aat gaa atg tta aaa tat tta aag aaa gat aga aat 2666Asp Ile Tyr Thr Asn Glu Met Leu Lys Tyr Leu Lys Lys Asp Arg Asn835 840 845ata ata ttc cta tct ccc gct atg tta gga gga tca gga ttg gtt aaa 2714Ile Ile Phe Leu Ser Pro Ala Met Leu Gly Gly Ser Gly Leu Val Lys850 855 860att agt gag cgt tat cca aat aat gta tat gat gta ggt ata gca gaa 2762Ile Ser Glu Arg Tyr Pro Asn Asn Val Tyr Asp Val Gly Ile Ala Glu865 870 875caa cat tct gta act ttc gca gca gct atg gca atg aat aag aaa tta 2810Gln His Ser Val Thr Phe Ala Ala Ala Met Ala Met Asn Lys Lys Leu880 885 890 895aaa ata caa tta tgt ata tat tcg acc ttt tta caa aga gca tat gat 2858Lys Ile Gln Leu Cys Ile Tyr Ser Thr Phe Leu Gln Arg Ala Tyr Asp900 905 910caa att ata cat gat ctt aat tta caa aat ata cct tta aag gtt ata 2906Gln Ile Ile His Asp Leu Asn Leu Gln Asn Ile Pro Leu Lys Val Ile915 920 925att gga aga agt gga tta gta gga gag gat ggg gca aca cat caa ggt 2954Ile Gly Arg Ser Gly Leu Val Gly Glu Asp Gly Ala Thr His Gln Gly930 935 940ata tat gat tta tct tat ctt ggg aca ctt aac aat gca tat ata ata 3002Ile Tyr Asp Leu Ser Tyr Leu Gly Thr Leu Asn Asn Ala Tyr Ile Ile945 950 955tct cca agt aat caa gtt gat ttg aaa aga gct ctt agg ttt gct tat 3050Ser Pro Ser Asn Gln Val Asp Leu Lys Arg Ala Leu Arg Phe Ala Tyr960 965 970 975tta gat aag gac cat tct gtg tat ata cgt ata ccc aga atg aac ata 3098Leu Asp Lys Asp His Ser Val Tyr Ile Arg Ile Pro Arg Met Asn Ile980 985 990tta agt gat aag tac atg aaa gga tat ttg aac att cat atg aaa aat 3146Leu Ser Asp Lys Tyr Met Lys Gly Tyr Leu Asn Ile His Met Lys Asn99510001005gag agc aaa aat atc gat gta aac gtg gat ata aac gat gat gta gat 3194Glu Ser Lys Asn Ile Asp Val Asn Val Asp Ile Asn Asp Asp Val Asp101010151020aaa tat agt gaa gaa tat atg gac gat gat aat ttt ata aaa tcg ttt 3242Lys Tyr Ser Glu Glu Tyr Met Asp Asp Asp Asn Phe Ile Lys Ser Phe102510301035att gga aaa tct aga att att aaa atg gat aat gaa aat aat aat aca 3290Ile Gly Lys Ser Arg Ile Ile Lys Met Asp Asn Glu Asn Asn Asn Thr1040 104510501055aat gaa cat tat tca agc aga gga gat aca cag aca aaa aaa aaa aaa 3338Asn Glu His Tyr Ser Ser Arg Gly Asp Thr Gln Thr Lys Lys Lys Lys106010651070gtt tgt atc ttt aac atg ggt agt atg ctt ttt aat gta att aat gct 3386Val Cys Ile Phe Asn Met Gly Ser Met Leu Phe Asn Val Ile Asn Ala107510801085ata aaa gaa att gaa aaa gaa caa tat att tca cat aat tat tct ttt 3434Ile Lys Glu Ile Glu Lys Glu Gln Tyr Ile Ser His Asn Tyr Ser Phe
109010951100tca att gtt gat atg ata ttt tta aat cct tta gat aaa aat atg ata 3482Ser Ile Val Asp Met Ile Phe Leu Asn Pro Leu Asp Lys Asn Met Ile110511101115gat cat gta ata aaa caa aat aaa cat caa tat tta att act tat gaa 3530Asp His Val Ile Lys Gln Asn Lys His Gln Tyr Leu Ile Thr Tyr Glu1120 112511301135gat aat act ata ggt ggt ttt tct aca cat ttc aat aat tat tta ata 3578Asp Asn Thr Ile Gly Gly Phe Ser Thr His Phe Asn Asn Tyr Leu Ile114011451150gaa aat aat tat att aca aaa cat aac tta tat gtt cat aat att tat 3626Glu Asn Asn Tyr Ile Thr Lys His Asn Leu Tyr Val His Asn Ile Tyr115511601165tta tct aat gag cca att gaa cat gca tct ttt aag gat caa caa gaa 3674Leu Ser Asn Glu Pro Ile Glu His Ala Ser Phe Lys Asp Gln Gln Glu117011751180gte gtc aaa atg gat aaa tgt agt ctt gtc aat aga att aaa aat tat 3722Val Val Lys Met Asp Lys Cys Ser Leu Val Asn Arg Ile Lys Asn Tyr118511901195ctt aaa aat aat cct aca tgatgtaaga taaatatata tttctaaaat 3770Leu Lys Asn Asn Pro Thr1200 1205tatttttttt ttatacttta atgtgtacaa taaaatatat atctaaatat attttatttg 3830tacgcttttt tttttttttt tttaattgtt atttttgtat at3872<210>4<211>1205<212>PRT<213>惡性瘧原蟲<400>4Met Ile Phe Asn Tyr Val Phe Phe Lys Asn Phe Val Pro Val Val Leu1 5 10 15Tyr Ile Leu Leu Ile Ile Tyr Ile Asn Leu Asn Gly Met Asn Asn Lys20 25 30Asn Gln Ile Lys Thr Glu Lys Ile Tyr Ile Lys Lys Leu Asn Arg Leu35 40 45Ser Arg Lys Asn Ser Leu Cys Ser Ser Lys Asn Lys Ile Ala Cys Leu20
50 55 60Phe Asp Ile GIy Asn Asp Asp Asn Arg Asn Thr Thr Tyr Gly Tyr Asn65 70 75 80Val Asn Val Lys Asn Asp Asp Ile Asn Ser Leu Leu Lys Asn Asn Tyr85 90 95Ser Asn Lys Leu Tyr Met Asp Lys Arg Lys Asn Ile Asn Asn Val Ile100 105 110Ser Thr Asn Lys Ile Ser Gly Ser Ile Ser Asn Ile Cys Ser Arg Asn115 120 125Gln Lys Glu Asn Glu Gln Lys Arg Asn Lys Gln Arg Cys Leu Thr Gln130 135 140Cys His Thr Tyr Asn Met Ser His Glu Gln Asp Lys Leu Ala Asn Asp145 150 155 160Asn Asn Arg Asn Asn Lys Lys Asn Phe Asn Leu Leu Phe Ile Asn Tyr165 170 175Phe Asn Leu Lys Arg Met Lys Asn Ser Leu Leu Asn Lys Asp Asn Phe180 185 190Phe Tyr Cys Lys Glu Lys Lys Leu Ser Phe Leu His Lys Ala Tyr Lys195 200 205Lys Lys Asn Cys Thr Phe Gln Asn Tyr Ser Leu Lys Arg Lys Ser Asn210 215 220Arg Asp Ser His Lys Leu Phe Ser Gly Glu Phe Asp Asp Tyr Thr Asn225 230 235 240Asn Asn Ala Leu Tyr Glu Ser Glu Lys Lys Glu Tyr Ile Thr Leu Asn245 250 255Asn Asn Asn Lys Asn Asn Asn Asn Lys Asn Asn Asp Asn Lys Asn Asn260 265 270Asp Asn Asn Asp Tyr Asn Asn Asn Asn Ser Cys Asn Asn Leu Gly Glu275 280 285Arg Ser Asn His Tyr Asp Asn Tyr Gly Gly Asp Asn Asn Asn Pro Cys290 295 300Asn Asn Asn Asn Asp Lys Tyr Asp Ile Gly Lys Tyr Phe Lys Gln Ile305 310 315 320Asn Thr Phe Ile Asn Ile Asp Glu Tyr Lys Thr Ile Tyr Gly Asp Glu
325 330 335Ile Tyr Lys Glu Ile Tyr Glu Leu Tyr Val Glu Arg Asn Ile Pro Glu340 345 350Tyr Tyr Glu Arg Lys Tyr Phe Ser Glu Asp Ile Lys Lys Ser Val Leu355 360 365Phe Asp Ile Asp Lys Tyr Asn Asp Val Glu Phe Glu Lys Ala Ile Lys370 375 380Glu Glu Phe Ile Asn Asn Gly Val Tyr Ile Asn Asn Ile Asp Asn Thr385 390 395 400Tyr Tyr Lys Lys Glu Asn Ile Leu Ile Met Lys Lys Ile Leu His Tyr405 410 415Phe Pro Leu Leu Lys Leu Ile Asn Asn Pro Ser Asp Leu Lys Lys Leu420 425 430Lys Lys Gln Tyr Leu Pro Leu Leu Ala His Glu Leu Lys Ile Phe Leu435 440 445Phe Phe Ile Val Asn Ile Thr Gly Gly His Phe Ser Ser Val Leu Ser450 455 460Ser Leu Glu Ile Gln Leu Leu Leu Leu Tyr Ile Phe Asn Gln Pro Tyr465 470 475 480Asp Asn Val Ile Tyr Asp Ile Gly His Gln Ala Tyr Val His Lys Ile485 490 495Leu Thr Gly Arg Lys Leu Leu Phe Leu Ser Leu Arg Asn Lys Lys Gly500 505 510Ile Ser Gly Phe Leu Asn Ile Phe Glu Ser Ile Tyr Asp Lys Phe Gly515 520 525Ala Gly His Ser Ser Thr Ser Leu Ser Ala Ile Gln Gly Tyr Tyr Glu530 535 540Ala Glu Trp Gln Val Lys Asn Lys Glu Lys Tyr Gly Asn Gly Asp Ile545 550 555 560Glu Ile Ser Asp Asn Ala Asn Val Thr Asn Asn Glu Arg Ile Phe Gln565 570 575Lys Gly Ile His Asn Asp Asn Asn Ile Asn Asn Asn Ile Asn Asn Asn580 585 590Asn Tyr Ile Asn Pro Ser Asp Val Val Gly Arg Glu Asn Thr Asn Val
595 600 605Pro Asn Val Arg Asn Asp Asn His Asn Val Asp Lys Val His Ile Ala610 615 620Ile Ile Gly Asp Gly Gly Leu Thr Gly Gly Met Ala Leu Glu Ala Leu625 630 635 640Asn Tyr Ile Ser Phe Leu Asn Ser Lys Ile Leu Ile Ile Tyr Asn Asp645 650 655Asn Gly Gln Val Ser Leu Pro Thr Asn Ala Val Ser Ile Ser Gly Asn660 665 670Arg Pro Ile Gly Ser Ile Ser Asp His Leu His Tyr Phe Val Ser Asn675 680 685Ile Glu Ala Asn Ala Gly Asp Asn Lys Leu Ser Lys Asn Ala Lys Glu690 695 700Asn Asn Ile Phe Glu Asn Leu Asn Tyr Asp Tyr Ile Gly Val Val Asn705 710 715 720Gly Asn Asn Thr Glu Glu Leu Phe Lys Val Leu Asn Asn Ile Lys Glu725 730 735Asn Lys Leu Lys Arg Ala Thr Val Leu His Val Arg Thr Lys Lys Ser740 745 750Asn Asp Phe Ile Asn Ser Lys Ser Pro Ile Ser Ile Leu His Ser Ile755 760 765Lys Lys Asn Glu Ile Phe Pro Phe Asp Thr Thr Ile Leu Asn Gly Asn770 775 780Ile His Lys Glu Asn Lys Ile Glu Glu Glu Lys Asn Val Ser Ser Ser785 790 795 800Thr Lys Tyr Asp Val Asn Asn Lys Asn Asn Lys Asn Asn Asp Asn Ser805 810 815Glu Ile Ile Lys Tyr Glu Asp Met Phe Ser Lys Glu Thr Phe Thr Asp820 825 830Ile Tyr Thr Asn Glu Met Leu Lys Tyr Leu Lys Lys Asp Arg Asn Ile835 840845Ile Phe Leu Ser Pro Ala Met Leu Gly Gly Ser Gly Leu Val Lys Ile850 855 860Ser Glu Arg Tyr Pro Asn Asn ValTyr Asp Val Gly Ile Ala Glu Gln865 870 875 880His Ser Val Thr Phe Ala Ala Ala Met Ala Met Asn Lys Lys Leu Lys885 890 895Ile Gln Leu Cys Ile Tyr Ser Thr Phe Leu Gln Arg Ala Tyr Asp Gln900 905 910Ile Ile His Asp Leu Asn Leu Gln Asn Ile Pro Leu Lys Val Ile Ile915 920 925Gly Arg Ser Gly Leu Val Gly Glu Asp Gly Ala Thr His Gln Gly Ile930 935 940Tyr Asp Leu Ser Tyr Leu Gly Thr Leu Asn Asn Ala Tyr Ile Ile Ser945 950 955 960Pro Ser Asn Gln Val Asp Leu Lys Arg Ala Leu Arg Phe Ala Tyr Leu965 970 975Asp Lys Asp His Ser Val Tyr Ile Arg Ile Pro Arg Met Asn Ile Leu980 985 990Ser Asp Lys Tyr Met Lys Gly Tyr Leu Asn Ile His Met Lys Asn Glu99510001005Ser Lys Asn Ile Asp Val Asn Val Asp Ile Asn Asp Asp Val Asp Lys101010151020Tyr Ser Glu Glu Tyr Met Asp Asp Asp Asn Phe Ile Lys Ser Phe Ile025103010351040Gly Lys Ser Arg Ile Ile Lys Met Asp Asn Glu Asn Asn Asn Thr Asn104510501055Glu His Tyr Ser Ser Arg Gly Asp Thr Gln Thr Lys Lys Lys Lys Val106010651070Cys Ile Phe Asn Met Gly Ser Met Leu Phe Asn Val Ile Asn Ala Ile107510801085Lys Glu Ile Glu Lys Glu Gln Tyr Ile Ser His Asn Tyr Ser Phe Ser109010951100Ile Val Asp Met Ile Phe Leu Asn Pro Leu Asp Lys Asn Met Ile Asp105111011151120His Val Ile Lys Gln Asn Lys His Gln Tyr Leu Ile Thr Tyr Glu Asp112511301135Asn Thr Ile Gly Gly Phe Ser Thr His Phe Asn Asn Tyr Leu Ile Glu
114011451150Asn Asn Tyr Ile Thr Lys His Asn Leu Tyr Val His Asn Ile Tyr Leu115511601165Ser Asn Glu Pro Ile Glu His Ala Ser Phe Lys Asp Gln Gln Glu Val117011751180Val Lys Met Asp Lys Cys Ser Leu Val Asn Arg Ile Lys Asn Tyr Leu185119011951200Lys Asn Asn Pro Thr1205<210>5<211>3147<212>DNA<213>惡性瘧原蟲<220><221>CDS<222>(199)..(2670)<400>5tttcattttt ctttacccac atatatatat atatatatat aatatatata tataatatta 60tatatttgat atatgattta aaattgtaac ataaaaaaaa taattatatt aaatatgtgt 120atacatctcc aacatataaa tattattttt tattattatt tttttttttt tttttcataa 180tgcctgaata accacaaa atg agt tat ata aaa aga ctg att ctt ttt atg 231Met Ser Tyr Ile Lys Arg Leu Ile Leu Phe Met1 5 10tta ctg ttt tat tct cat gta aaa att aaa aaa tta ttt att aaa att 279Leu Leu Phe Tyr Ser His Val Lys Ile Lys Lys Leu Phe Ile Lys Ile15 20 25tct aat gta aac ata ttt ttt gca gaa gca aag aaa aat gga aaa aag 327Ser Asn Val Asn Ile Phe Phe Ala Glu Ala Lys Lys Asn Gly Lys Lys30 35 40gaa ttc ttt ctt ttt tta cta aat ata aaa aaa aat agc caa cag aaa 375Glu Phe Phe Leu Phe Leu Leu Asn Ile Lys Lys Asn Ser Gln Gln Lys45 50 55aaa act tat cat att acc aaa agg aat acc ata aat aaa agt gat ttt 423Lys Thr Tyr His Ile Thr Lys Arg Asn Thr Ile Asn Lys Ser Asp Phe60 65 70 75tta tat tct tta cta aat gaa gaa ggg aat tct tca aaa aag gaa tat 471Leu Tyr Ser Leu Leu Asn Glu Glu Gly Asn Ser Ser Lys Lys Glu Tyr80 85 90aaa aat tta aaa gat gaa gaa aaa tat aat atc ata caa aat ata aaa 519Lys Asn Leu Lys Asp Glu Glu Lys Tyr Asn Ile Ile Gln Asn Ile Lys95 100 105aaa tat tgt gaa tgt act aaa aaa tat aaa agg ctc cca aca cga gaa 567Lys Tyr Cys Glu Cys Thr Lys Lys Tyr Lys Arg Leu Pro Thr Arg Glu110 115 120gta gtt att gga aat gtt aaa att gga gga aat aat aaa ata gct att 615Val Val Ile Gly Asn Val Lys Ile Gly Gly Asn Asn Lys Ile Ala Ile125 130 135caa act atg gct agc tgt gat aca aga aat gta gaa gaa tgt gta tat 663Gln Thr Met Ala Ser Cys Asp Thr Arg Asn Val Glu Glu Cys Val Tyr140 145 150 155caa att aga aaa tgt aaa gat ttg ggt gct gac att gta agg ttg act 711Gln Ile Arg Lys Cys Lys Asp Leu Gly Ala Asp Ile Val Arg Leu Thr160 165 170gtt caa gga gtt caa gaa gca caa gct agt tat cat att aaa gaa aaa 759Val Gln Gly Val Gln Glu Ala Gln Ala Ser Tyr His Ile Lys Glu Lys175 180 185tta tta tct gaa aat gta aat atc cca tta gta gca gat att cat ttt 807Leu Leu Ser Glu Asn Val Asn Ile Pro Leu Val Ala Asp Ile His Phe190 195 200aat cct aaa ata gct tta atg gca gct gat gtg ttt gaa aaa att cga 855Asn Pro Lys Ile Ala Leu Met Ala Ala Asp Val Phe Glu Lys Ile Arg205 210 215gtg aat cca gga aat tat gtt gat gga aga aaa aaa tgg ata gat aaa 903Val Asn Pro Gly Asn Tyr Val Asp Gly Arg Lys Lys Trp Ile Asp Lys220 225 230 235gtt tat aaa act aaa gaa gaa ttt gat gaa ggg aaa tta ttt ata aaa 951Val Tyr Lys Thr Lys Glu Glu Phe Asp Glu Gly Lys Leu Phe Ile Lys240 245 250gaa aaa ttt gta cca tta att gaa aaa tgt aaa aga tta aat aga gca 999Glu Lys Phe Val Pro Leu Ile Glu Lys Cys Lys Arg Leu Asn Arg Ala255 260 265ata aga att gga aca aat cat gga tcc ctt tca tct cga gta tta tca 1047Ile Arg Ile Gly Thr Asn His Gly Ser Leu Ser Ser Arg Val Leu Ser
270 275 280tat tat gga gat aca cca tta ggt atg gta gaa tcg gct ttt gag ttt 1095Tyr Tyr Gly Asp Thr Pro Leu Gly Met Val Glu Ser Ala Phe Glu Phe285 290 295tct gat tta tgt att gaa aac aat ttt tac aat ctt gtt ttt tct atg 1143Ser Asp Leu Cys Ile Glu Asn Asn Phe Tyr Asn Leu Val Phe Ser Met300 305 310 315aaa gct tct aat gct tat gtt atg ata caa tct tat aga tta tta gta 1191Lys Ala Ser Asn Ala Tyr Val Met Ile Gln Ser Tyr Arg Leu Leu Val320 325 330tct aaa caa tat gaa aga aat atg atg ttc cct ata cat tta gga gtt 1239Ser Lys Gln Tyr Glu Arg Asn Met Met Phe Pro Ile His Leu Gly Val335 340 345aca gaa gca gga ttt ggt gat aat gga aga ata aaa tct tat tta ggt 1287Thr Glu Ala Gly Phe Gly Asp Asn Gly Arg Ile Lys Ser Tyr Leu Gly350 355 360ata gga tct tta tta tat gat ggt ata gga gat acc att cgt ata tcc 1335Ile Gly Ser Leu Leu Tyr Asp Gly Ile Gly Asp Thr Ile Arg Ile Ser365 370 375tta aca gaa gat cct tgg gaa gag tta act cct tgt aaa aaa tta gtt 1383Leu Thr Glu Asp Pro Trp Glu Glu Leu Thr Pro Cys Lys Lys Leu Val380 385 390 395gaa aat tta aag aaa aga ata ttt tat aat gaa aat ttt aaa gaa gat 1431Glu Asn Leu Lys Lys Arg Ile Phe Tyr Asn Glu Asn Phe Lys Glu Asp400 405 410aat gaa tta aaa aat aat gaa atg gat acc aaa aat cta tta aat ttt 1479Asn Glu Leu Lys Asn Asn Glu Met Asp Thr Lys Asn Leu Leu Asn Phe415 420 425gaa gaa aat tat cga aat ttt aat aat ata aaa aaa aga aat gta gaa 1527Glu Glu Asn Tyr Arg Asn Phe Asn Asn Ile Lys Lys Arg Asn Val Glu430 435 440aaa aat aat aat gta tta cat gaa gag tgc act ata ggt aat gta gta 1575Lys Asn Asn Asn Val Leu His Glu Glu Cys Thr Ile Gly Asn Val Val445 450 455acc ata aaa gag tta gaa gat tct ctg caa att ttt aaa gat tta aat 1623Thr Ile Lys Glu Leu Glu Asp Ser Leu Gln Ile Phe Lys Asp Leu Asn460 465 470 475tta gaa gta gat tca aat gga aat ttg aaa aag gga gcc aaa aca act 1671Leu Glu Val Asp Ser Asn Gly Asn Leu Lys Lys Gly Ala Lys Thr Thr480 485 490gat atg gtt att ata aat gat ttt cat aat ata aca aat tta gga aaa 1719Asp Met Val Ile Ile Asn Asp Phe His Asn Ile Thr Asn Leu Gly Lys495 500 505aaa act gtg gat aaa tta atg caa gtg gga att aat ata gta gtt caa 1767Lys Thr Val Asp Lys Leu Met Gln Val Gly Ile Asn Ile Val Val Gln510 515 520tat gaa cca cat aat ata gaa ttt ata gaa aaa atg gaa cca aat aat 1815Tyr Glu Pro His Asn Ile Glu Phe Ile Glu Lys Met Glu Pro Asn Asn525 530 535gat aat aat aat aat aat aat aat aat aat ata tta ttt tat gtg gat 1863Asp Asn Asn Asn Asn Asn Asn Asn Asn Asn Ile Leu Phe Tyr Val Asp540 545 550 555ata aaa aat att atg aac agt tca gaa aaa aat att aaa tta agt aat 1911Ile Lys Asn Ile Met Asn Ser Ser Glu Lys Asn Ile Lys Leu Ser Asn560 565 570tct aaa gga tat gga tta att tta aac gga aaa gaa gat ata caa acc 1959Ser Lys Gly Tyr Gly Leu Ile Leu Asn Gly Lys Glu Asp Ile Gln Thr575 580 585ata aaa aaa ata aaa gaa tta aat cgt cgt cct tta ttc att cta tta 2007Ile Lys Lys Ile Lys Glu Leu Asn Arg Arg Pro Leu Phe Ile Leu Leu590 595 600aaa tca gat aac ata tat gaa cat gta tta ata acc aga aga att aat 2055Lys Ser Asp Asn Ile Tyr Glu His Val Leu Ile Thr Arg Arg Ile Asn605 610 615gaa ctt tta caa tcc tta aat ata aat ata cct tat ata cat tat gtt 2103Glu Leu Leu Gln Ser Leu Asn Ile Asn Ile Pro Tyr Ile His Tyr Val620 625 630 635gat att aat tca aac aat tat gat gat ata tta gtt aat tca aca tta 2151Asp Ile Asn Ser Asn Asn Tyr Asp Asp Ile Leu Val Asn Ser Thr Leu640 645 650tat gca gga agt tgt ttg atg gat tta atg ggg gat ggt ctt att gtt 2199Tyr Ala Gly Ser Cys Leu Met Asp Leu Met Gly Asp Gly Leu Ile Val655 660 665aac gta act aat gat gtt ctt aca aat aaa aaa aag ata gaa aca aaa 2247Asn Val Thr Asn Asp Val Leu Thr Asn Lys Lys Lys Ile Glu Thr Lys670 675 680tat gat gaa aaa gaa gaa gta gag gaa gag gga aac aat aaa gat att 2295Tyr Asp Glu Lys Glu Glu Val Glu Glu Glu Gly Asn Asn Lys Asp Ile685 690 695cat aga ctt ttg agc aga gtt gca tta aat tca ttt tta aca tta aat 2343His Arg Leu Leu Ser Arg Val Ala Leu Asn Ser Phe Leu Thr Leu Asn700 705 710 715att tta caa gat aca aga ata cgt tta ttt aaa aca gat tat ata gcc 2391Ile Leu Gln Asp Thr Arg Ile Arg Leu Phe Lys Thr Asp Tyr Ile Ala720 725 730tgc cca tct tgt gga aga act tta ttt aat ata caa gaa act act aaa 2439Cys Pro Ser Cys Gly Arg Thr Leu Phe Asn Ile Gln Glu Thr Thr Lys735 740 745aaa att atg aaa tta aca ggg cac tta aaa ggc gtt aaa att gca gtc 2487Lys Ile Met Lys Leu Thr Gly His Leu Lys Gly Val Lys Ile Ala Val750 755 760atg gga tgt att gtt aat ggt ata gga gaa atg gca gat gca cat ttt 2535Met Gly Cys Ile Val Asn Gly Ile Gly Glu Met Ala Asp Ala His Phe765 770 775ggt tat gtt ggt agt gca cct aaa aaa att gat tta tat tat ggt aaa 2583Gly Tyr Val Gly Ser Ala Pro Lys Lys Ile Asp Leu Tyr Tyr Gly Lys780 785 790 795gag tta gta gaa aga aat ata cct gag gaa gaa gct tgt gat aaa ttg 2631Glu Leu Val Glu Arg Asn Ile Pro Glu Glu Glu Ala Cys Asp Lys Leu800 805 810ata gaa tta att aaa aaa cat aac aaa tgg aaa gat cca taaattgaat2680Ile Glu Leu Ile Lys Lys His Asn Lys Trp Lys Asp Pro815 820atggacaagt atttatttat ttatttatct tatatataat atattataaa tttttcgatg 2740tatttttcct tttaaaattt tatttttttt ttattttttt ttttgaagta atatttataa 2800tgcatacata atattaaaat gtgtattata taataatatc attttattgt tattttaaaa 2860gactaatacc aagaacaatt ttttaataat cattcttata acttgttaaa tatatatata 2920tatatatata tatttattta tttatattta tatttattta tttttggtat atgaaaagta 2980aaaatataat aatttaaaag tatttacaaa ataaataata ttatatatct gtttttatat 3040atatgttaat ggaaaaggag aaaataaata aataaaacaa acaaaataac atatatatat 3100atatatatat actgaatgag aaagaaaaaa aaaagaaaag gatacga 3147<210>6<211>824<212>PRT<213>惡性瘧原蟲<400>6Met Ser Tyr Ile Lys Arg Leu Ile Leu Phe Met Leu Leu Phe Tyr Ser1 5 10 15His Val Lys Ile Lys Lys Leu Phe Ile Lys Ile Ser Asn Val Asn Ile20 25 30Phe Phe Ala Glu Ala Lys Lys Asn Gly Lys Lys Glu Phe Phe Leu Phe35 40 45Leu Leu Asn Ile Lys Lys Asn Ser Gln Gln Lys Lys Thr Tyr His Ile50 55 60Thr Lys Arg Asn Thr Ile Asn Lys Ser Asp Phe Leu Tyr Ser Leu Leu65 70 75 80Asn Glu Glu Gly Asn Ser Ser Lys Lys Glu Tyr Lys Asn Leu Lys Asp85 90 95Glu Glu Lys Tyr Asn Ile Ile Gln Asn Ile Lys Lys Tyr Cys Glu Cys100 105 110Thr Lys Lys Tyr Lys Arg Leu Pro Thr Arg Glu Val Val Ile G1y Asn115 120 125Val Lys Ile Gly Gly Asn Asn Lys Ile Ala Ile Gln Thr Met Ala Ser130 135 140Cys Asp Thr Arg Asn Val Glu Glu Cys Val Tyr Gln Ile Arg Lys Cys145 150 155 160Lys Asp Leu Gly Ala Asp Ile Val Arg Leu Thr Val Gln G1y Val Gln165 170 175Glu Ala Gln Ala Ser Tyr His Ile Lys Glu Lys Leu Leu Ser Glu Asn180 185 190Val Asn Ile Pro Leu Val Ala Asp Ile His Phe Asn Pro Lys Ile Ala195 200 205Leu Met Ala Ala Asp Val Phe Glu Lys Ile Arg Val Asn Pro Gly Asn210 215 220Tyr Val Asp Gly Arg Lys Lys Trp Ile Asp Lys Val Tyr Lys Thr Lys225 230 235 240Glu Glu Phe Asp Glu Gly Lys Leu Phe Ile Lys Glu Lys Phe Val Pro245 250 255Leu Ile Glu Lys Cys Lys Arg Leu Asn Arg Ala Ile Arg Ile Gly Thr260 265 270Asn His Gly Ser Leu Ser Ser Arg Val Leu Ser Tyr Tyr Gly Asp Thr275 280 285Pro Leu Gly Met Val Glu Ser Ala Phe Glu Phe Ser Asp Leu Cys Ile290 295 300Glu Asn Asn Phe Tyr Asn Leu Val Phe Ser Met Lys Ala Ser Asn Ala305 310 315 320Tyr Val Met Ile Gln Ser Tyr Arg Leu Leu Val Ser Lys Gln Tyr Glu325 330 335Arg Asn Met Met Phe Pro Ile His Leu Gly Val Thr Glu Ala Gly Phe340 345 350Gly Asp Asn Gly Arg Ile Lys Ser Tyr Leu Gly Ile Gly Ser Leu Leu355 360 365Tyr Asp Gly Ile Gly Asp Thr Ile Arg Ile Ser Leu Thr Glu Asp Pro370 375 380Trp Glu Glu Leu Thr Pro Cys Lys Lys Leu Val Glu Asn Leu Lys Lys385 390 395 400Arg Ile Phe Tyr Asn Glu Asn Phe Lys Glu Asp Asn Glu Leu Lys Asn405 410 415Asn Glu Met Asp Thr Lys Asn Leu Leu Asn Phe Glu Glu Asn Tyr Arg420 425 430Asn Phe Asn Asn Ile Lys Lys Arg Asn Val Glu Lys Asn Asn Asn Val435 440 445Leu His Glu Glu Cys Thr Ile Gly Asn Val Val Thr Ile Lys Glu Leu450 455 460Glu Asp Ser Leu Gln Ile Phe Lys Asp Leu Asn Leu Glu Val Asp Ser465 470 475 480Asn Gly Asn Leu Lys Lys Gly Ala Lys Thr Thr Asp Met Val Ile Ile485 490 495Asn Asp Phe His Asn Ile Thr Asn Leu Gly Lys Lys Thr Val Asp Lys
500 505 510Leu Met Gln Val Gly lle Asn Ile Val Val Gln Tyr Glu Pro His Asn515 520 525Ile Glu Phe Ile Glu Lys Met Glu Pro Asn Asn Asp Asn Asn Asn Asn530 535 540Asn Asn Asn Asn Asn Ile Leu Phe Tyr Val Asp Ile Lys Asn Ile Met545 550 555 560Asn Ser Ser Glu Lys Asn Ile Lys Leu Ser Asn Ser Lys Gly Tyr Gly565 570 575Leu Ile Leu Asn Gly Lys Glu Asp Ile Gln Thr Ile Lys Lys Ile Lys580 585 590Glu Leu Asn Arg Arg Pro Leu Phe Ile Leu Leu Lys Ser Asp Asn Ile595 600 605Tyr Glu His Val Leu Ile Thr Arg Arg Ile Asn Glu Leu Leu Gln Ser610 615 620Leu Asn Ile Asn Ile Pro Tyr Ile His Tyr Val Asp Ile Asn Ser Asn625 630 635 640Asn Tyr Asp Asp Ile Leu Val Asn Ser Thr Leu Tyr Ala Gly Ser Cys645 650 655Leu Met Asp Leu Met Gly Asp Gly Leu Ile Val Asn Val Thr Asn Asp660 665 670Val Leu Thr Asn Lys Lys Lys Ile Glu Thr Lys Tyr Asp Glu Lys Glu675 680 685Glu Val Glu Glu Glu Gly Asn Asn Lys Asp Ile His Arg Leu Leu Ser690 695 700Arg Val Ala Leu Asn Ser Phe Leu Thr Leu Asn Ile Leu Gln Asp Thr705 710 715 720Arg Ile Arg Leu Phe Lys Thr Asp Tyr Ile Ala Cys Pro Ser Cys Gly725 730 735Arg Thr Leu Phe Asn Ile Gln Glu Thr Thr Lys Lys Ile Met Lys Leu740 745 750Thr Gly His Leu Lys Gly Val Lys Ile Ala Val Met Gly Cys Ile Val755 760 765Asn Gly Ile Gly Glu Met Ala Asp Ala His Phe Gly Tyr Val Gly Ser
770 775 780Ala Pro Lys Lys Ile Asp Leu Tyr Tyr Gly Lys Glu Leu Val Glu Arg785 790 795 800Asn Ile Pro Glu Glu Glu Ala Cys Asp Lys Leu Ile Glu Leu Ile Lys805 810 815Lys His Asn Lys Trp Lys Asp Pro820
權(quán)利要求
1．DNA序列，該序列編碼具有SEQ ID No.2顯示的氨基酸序列的多肽或編碼SEQ ID No.2的多肽的類似物或者衍生物的DNA序列，該類似物或衍生物是其中一個或多個氨基酸缺失、添加或被其它氨基酸替代。
2．DNA序列，該序列編碼具有SEQ ID No.4顯示的氨基酸序列的多肽或編碼SEQ ID No.4的多肽的類似物或者衍生物的DNA序列，該類似物或衍生物是其中一個或多個氨基酸缺失、添加或被其它氨基酸替代。
3．DNA序列，該序列編碼具有SEQ ID No.6顯示的氨基酸序列的多肽或編碼SEQ ID No.6的多肽的類似物或者衍生物的DNA序列，該類似物或衍生物是其中一個或多個氨基酸缺失、添加或被其它氨基酸替代。
4．根據(jù)權(quán)利要求1-3任一項所述的DNA序列，特征在于它還包括功能調(diào)節(jié)信號，特別是啟動子、操縱基因、增強(qiáng)子、核糖體結(jié)合位點(diǎn)。
5．具有下面的亞序列的DNA序列ⅰ)在病毒、真核生物和原核生物中具有活性并且確保在預(yù)定的靶組織或靶細(xì)胞中形成RNA的啟動子，ⅱ)根據(jù)權(quán)利要求1-3任一項所述的DNA序列，ⅲ)在病毒、真核生物和原核生物中導(dǎo)致聚(A)殘基加入到RNA的3’末端的3’未翻譯序列。
6．生產(chǎn)轉(zhuǎn)基因病毒、真核生物和原核生物以改變類異戊二烯含量的方法，特征在于使用或不用載體將權(quán)利要求4或5的DNA序列轉(zhuǎn)移和摻入到病毒、真核和原核細(xì)胞的基因組中。
7．轉(zhuǎn)基因系統(tǒng)，特別是植物和植物細(xì)胞，所述植物和植物細(xì)胞含有一個或多個權(quán)利要求1-5的DNA序列作為“外源”或“附加”DNA，該序列被表達(dá)。
8．含有一個或多個權(quán)利要求1-5的DNA序列的表達(dá)載體。
9．蛋白質(zhì)，它參與1-脫氧-D-木酮糖-5-磷酸代謝途徑并且a)由SEQ ID No.1,3或5的DNA序列編碼或b)由與SEQ ID No.1、3、5的DNA序列或這些DNA序列的片段在編碼成熟蛋白質(zhì)的DNA區(qū)域雜交的DNA序列編碼。
10．根據(jù)權(quán)利要求9所述的蛋白質(zhì)，是可從寄生蟲的培養(yǎng)物上清液或從破裂的寄生蟲獲得的并且通過層析和電泳方法純化。
11．根據(jù)權(quán)利要求9和10之一所述的蛋白質(zhì)，特征在于它a)是外源DNA的病毒、原核生物或真核生物的表達(dá)產(chǎn)物，b)是由SEQ IDNo.1、3或5的序列編碼的或由與SEQ ID No.1、3、5的DNA序列或這些DNA序列的片段在編碼成熟蛋白質(zhì)的DNA區(qū)域雜交的DNA序列編碼的，或c)是由在沒有遺傳密碼的簡并時與b)中定義的序列雜交的并且編碼具有相應(yīng)的氨基酸序列的多肽的DNA序列編碼的。
12．根據(jù)前面任一項權(quán)利要求所述的蛋白質(zhì)，特征在于它包括SEQ ID No.2、4或6的氨基酸序列。
13．用于測定gcpE蛋白的酶促活性的方法，特征在于檢測糖或磷糖或類異戊二烯生物合成的前體的磷酸化，特別是2-C-甲基-D-赤蘚糖醇、2-C-甲基-D-赤蘚糖醇磷酸酯的磷酸化，特別是2-C-甲基-D-赤蘚糖醇-4-磷酸酯、2-C-甲基-D-赤蘚糖、2-C-甲基-D-赤蘚糖磷酸酯的磷酸化，特別是2-C-甲基-D-赤蘚糖-4-磷酸酯的磷酸化，以及磷酸酯和醇前體的磷酸化。
14．根據(jù)權(quán)利要求13所述的方法，特征在于檢測下面磷酸酯或醇的磷酸化CH2(OH)-C(CH3)=C(OH)-CH2-O-PO(OH)2,CH2(OH)-C(CH3)=C(OH)-CH2-OH,CH2(OH)-CH(CH3)-CO-CH2-O-PO(OH)2,CH2(OH)-CH(CH3)-CO-CH2OH,CH2=C(CH3)-CO-CH2-O-PO(OH)2,CH2=C(CH3)-CO-CH2-OH,CH2=C(CH3)-CH(OH)-CH2-O-PO(OH)2,CH2=C(CH3)-CH(OH)-CH2-OH,CH2(OH)-C(=CH2)-C(OH)-CH2-O-PO(OH)2,CH2(OH)-C(=CH2)-C(OH)-CH2-OH,CHO-CH(CH3)-CH(OH)-CH2-O-PO-(OH)2,CHO-CH(CH3)-CH(OH)-CH2-OH,CH2(OH)-C(OH)(CH3)-CH=CH-O-PO(OH)2,CH2(OH)-C(OH)(CH3)-CH=CH-OHCH(OH)=C(CH3)-CH(OH)-CH2-O-PO(OH)2,CH(OH)=C(CH3)-CH(OH)-CH2-OH(CH3)2HC-CO-CH2-O-PO(OH)2,(CH3)2HC-CO-CH2-O-H,(CH3)2HC-CH(OH)-CH2-O-PO(OH)2,(CH3)2HC-CH(OH)-CH2-O-H。
15．DOXP合酶和DOXP還原異構(gòu)酶的酶促活性的組合測定方法，特征在于檢測甘油醛-3-磷酸到2-C-甲基赤蘚糖醇-4-磷酸酯的轉(zhuǎn)變。
16．篩選用于治療人類和動物的感染過程的化合物的方法，其中該方法包括a)提供含有重組表達(dá)載體的宿主細(xì)胞，其中載體包括至少SEQ ID No.1,SEQ ID No.3或SEQ ID No.5的寡核苷酸序列或其變體或同系物的一部分，另外提供被懷疑對人和動物具有抗真菌、抗菌、抗寄生蟲或抗病毒的作用的化合物，b)將宿主細(xì)胞與該化合物接觸，以及c)測定該化合物的抗微生物、抗真菌、抗菌、抗寄生蟲或抗病毒作用。
17.篩選用于治療植物的化合物的方法，其中該方法包括a)提供含有重組表達(dá)載體的宿主細(xì)胞，其中載體包括至少SEQ ID No.1、SEQ ID No.3或SEQ ID No.5的寡核苷酸序列或其變體或同系物的一部分，另外提供被懷疑對植物具有抗微生物、抗病毒、抗寄生蟲、殺細(xì)菌、殺真菌或除草的作用的化合物，b)將宿主細(xì)胞與該化合物接觸，以及c)測定該化合物的抗微生物、抗病毒、抗寄生蟲、殺細(xì)菌、殺真菌或除草的作用。
18．權(quán)利要求1-5任一項的DNA或權(quán)利要求9-12任一項的蛋白質(zhì)或權(quán)利要求7的轉(zhuǎn)基因系統(tǒng)在預(yù)防或治療人類和動物的疾病中的用途。
全文摘要
本發(fā)明涉及1－脫氧－D－木酮糖－5－磷酸還原異構(gòu)酶基因,1－脫氧－D－木酮糖－5－磷酸合酶基因以及1－脫氧－D－木酮糖生物合成途徑的gcpE基因,并涉及它們在轉(zhuǎn)化載體、宿主生物體和植物以及測定抑制這一生物合成途徑的物質(zhì)中的用途。
文檔編號A61P31/04GK1319134SQ99811200
公開日2001年10月24日 申請日期1999年9月22日 優(yōu)先權(quán)日1998年9月22日
發(fā)明者哈?！ぶ祚R 申請人:朱馬制藥有限公司