CAJ | 학술논문

对鹅细小病毒YZ株非结构蛋白1的基因进行克隆、测序、比对及蛋白质结构和功能等生物信息学分析,探究了非结构蛋白1的基因分子特征和理化特性及病理学相关致病机理。通过PCR、质粒克隆及核苷酸序列测定方法获得病毒YZ株非结构蛋白1基因序列,采用DNAsis、Mega 5.10、Blast等多种生物学软件完成核苷酸序列及氨基酸序列和系统进化分析。结果显示,鹅细小病毒YZ株非结构蛋白1的基因全长1881个核苷酸,编码627个氨基酸。与GPV-GB标准株、MDPV、ChPV、AAV的NS1基因序列存在很高的同源相似性,与MDPV的亲缘关系最近,提示番鸭和鹅细小病毒来自共同的祖先,其宿主的不同而演化为不同的分支。对应GPVNS1-YZ的同源区段序列(293~524)含231个氨基酸,分子量为26.446 kDa、PI=6.04,带负电的氨基酸30个,带正电的氨基酸28个,体外表达半衰期为5.5 h时,不稳定系数为40.23,脂肪系数为78.01,平均疏水值为-0.43,表明这一蛋白质为亲水性蛋白；该蛋白还存在9个抗原肽的位置：55~69,185~195,94~129,156~167,19~30,76~83,6~12,205~210,40~45,出现16个可能的B细胞抗原表位,表明它具有较高的免疫原性；但不存在跨膜片段,也不含信号肽和二硫键,可能不会分泌到宿主细胞的细胞膜外发生作用；其二级结构包含α-螺旋占30.74%,β-折叠占17.32%,无规则卷曲占51.95%,三级结构近似球状,包含一个SF3解旋酶超家族的DNA病毒解旋酶功能域( AA17-AA173),提示鹅细小病毒的非结构蛋白1可能在感染发生后参与病毒DNA自我复制中的DNA解旋过程。
대아세소병독YZ주비결구단백1적기인진행극륭、측서、비대급단백질결구화공능등생물신식학분석,탐구료비결구단백1적기인분자특정화이화특성급병이학상관치병궤리。통과PCR、질립극륭급핵감산서렬측정방법획득병독YZ주비결구단백1기인서렬,채용DNAsis、Mega 5.10、Blast등다충생물학연건완성핵감산서렬급안기산서렬화계통진화분석。결과현시,아세소병독YZ주비결구단백1적기인전장1881개핵감산,편마627개안기산。여GPV-GB표준주、MDPV、ChPV、AAV적NS1기인서렬존재흔고적동원상사성,여MDPV적친연관계최근,제시번압화아세소병독래자공동적조선,기숙주적불동이연화위불동적분지。대응GPVNS1-YZ적동원구단서렬(293~524)함231개안기산,분자량위26.446 kDa、PI=6.04,대부전적안기산30개,대정전적안기산28개,체외표체반쇠기위5.5 h시,불은정계수위40.23,지방계수위78.01,평균소수치위-0.43,표명저일단백질위친수성단백；해단백환존재9개항원태적위치：55~69,185~195,94~129,156~167,19~30,76~83,6~12,205~210,40~45,출현16개가능적B세포항원표위,표명타구유교고적면역원성；단불존재과막편단,야불함신호태화이류건,가능불회분비도숙주세포적세포막외발생작용；기이급결구포함α-라선점30.74%,β-절첩점17.32%,무규칙권곡점51.95%,삼급결구근사구상,포함일개SF3해선매초가족적DNA병독해선매공능역( AA17-AA173),제시아세소병독적비결구단백1가능재감염발생후삼여병독DNA자아복제중적DNA해선과정。
To conduct cloning and sequencing of non structural protein 1 ( NS1 ) gene from goose parvovirus YZ strain(GPV YZ) and make sequence alignment and analysis of protein structure and function by bioinformatics,and research the characteristics of NS1 gene and the physicochemical properties and pathogenic mechanism of NS1 pro-tein. The sequences of GPVNS1-YZ gene were obtained by PCR, plasmid cloning and sequencing. Bioinformatics was used to analyze the nucleic acid data,deduced amino acid sequence and phylogenetic trees. The result of se-quence analysis showed that the gene of GPVNS1-YZ protein was 1 881 nt in length,which coded 627 amino acids polyprotein. There was high homology in NS1 gene sequences and the phylogenetic relationships of MDPV recently, when compared with selected GPV-GB,MDPV,ChPV and AAV strains in GenBank. It therefore suggested MDPV and GPV from a common ancestor,the host and different evolution for different branch. The homology segment se-quences corresponding to GPVNS1-YZ contained 231 amino acids and the molecular weight,PI,positively or nega-tively charged amino acid, estimated half-life, instability index, aliphatic index and average hydrophobicity were 26. 446 kDa,6. 04,30,28,5. 5 h,40. 23,78. 01,-0. 43 respectively,and indicated that the protein was a hydro-philic protein. There were nine antigenic peptides and 16 B cell epitopes in the protein respectively,which showed high immunogenicity. However,there existed no transmembrane domain,signal peptide and two disulfide bonds in the protein ,and it might not be secreted into the extracellular membrane of host cell for function. The protein struc-ture study indicated that alpha-helix,beta-sheet,random coil were 30. 74%,17. 32%,51. 95% respectively. There was the virus DNA helicase domain of SF3 helicase superfamily in the tertiary structure. This study indicated that GPV NS1 protein might participate in DNA unwinding process during viral DNA replication,when infection occurs.