南方医科大学学报
南方醫科大學學報
남방의과대학학보
JOURNAL OF SOUTHERN MEDICAL UNIVERSITY
2009年
12期
2371-2374
,共4页
结核分枝杆菌%毒力分泌系统%Rv3871%致病性%生物信息学
結覈分枝桿菌%毒力分泌繫統%Rv3871%緻病性%生物信息學
결핵분지간균%독력분비계통%Rv3871%치병성%생물신식학
tuberculosis%virulent protein secretion system%Rv3871%pathogenicity%bioinformatics
目的 对结核分枝杆菌H37Rv毒株RD1毒力分泌系统Rv3871基因进行克隆和蛋白表达,运用生物信息学分析该基因在结核杆菌毒力蛋白分泌过程中的作用.方法 以结核分枝杆菌H37Rv基因组为模板,以PCR扩增出完整的Rv3871基因,将其插入原核表达载体pET32a(+),对其核苷酸序列进行测定.利用生物信息学软件对结核杆菌Rv3871进行结构功能分析并与多种分枝杆菌进行同源性比对.结果 经分子克隆的Rv3871酶切片段与预期大小符合,基因序列与Genbank报道一致,SDS-PAGE检测到相对分子质量为84 000的特异性表达蛋白,生物信息学分析发现两个FtsK/SpoEⅢ样结构域,同源性比对则发现在致病性分枝杆菌与卡介苗等非致病菌Rv3971基因对应区域的结构完整程度有较为显著的差别.结论 本研究通过对结核分枝杆菌RV3871基因进行分子克隆,特异蛋白表达和基因序列测定,提供了该基因的结构功能特征,并通过生物信息学与同源性对比分析,发现Rv3871基因在致病和非致病分枝杆菌毒力分泌作用中的结构和功能差异,为结核病发病机制阐明及其治疗药靶的筛选提供理论和实验依据.
目的 對結覈分枝桿菌H37Rv毒株RD1毒力分泌繫統Rv3871基因進行剋隆和蛋白錶達,運用生物信息學分析該基因在結覈桿菌毒力蛋白分泌過程中的作用.方法 以結覈分枝桿菌H37Rv基因組為模闆,以PCR擴增齣完整的Rv3871基因,將其插入原覈錶達載體pET32a(+),對其覈苷痠序列進行測定.利用生物信息學軟件對結覈桿菌Rv3871進行結構功能分析併與多種分枝桿菌進行同源性比對.結果 經分子剋隆的Rv3871酶切片段與預期大小符閤,基因序列與Genbank報道一緻,SDS-PAGE檢測到相對分子質量為84 000的特異性錶達蛋白,生物信息學分析髮現兩箇FtsK/SpoEⅢ樣結構域,同源性比對則髮現在緻病性分枝桿菌與卡介苗等非緻病菌Rv3971基因對應區域的結構完整程度有較為顯著的差彆.結論 本研究通過對結覈分枝桿菌RV3871基因進行分子剋隆,特異蛋白錶達和基因序列測定,提供瞭該基因的結構功能特徵,併通過生物信息學與同源性對比分析,髮現Rv3871基因在緻病和非緻病分枝桿菌毒力分泌作用中的結構和功能差異,為結覈病髮病機製闡明及其治療藥靶的篩選提供理論和實驗依據.
목적 대결핵분지간균H37Rv독주RD1독력분비계통Rv3871기인진행극륭화단백표체,운용생물신식학분석해기인재결핵간균독력단백분비과정중적작용.방법 이결핵분지간균H37Rv기인조위모판,이PCR확증출완정적Rv3871기인,장기삽입원핵표체재체pET32a(+),대기핵감산서렬진행측정.이용생물신식학연건대결핵간균Rv3871진행결구공능분석병여다충분지간균진행동원성비대.결과 경분자극륭적Rv3871매절편단여예기대소부합,기인서렬여Genbank보도일치,SDS-PAGE검측도상대분자질량위84 000적특이성표체단백,생물신식학분석발현량개FtsK/SpoEⅢ양결구역,동원성비대칙발현재치병성분지간균여잡개묘등비치병균Rv3971기인대응구역적결구완정정도유교위현저적차별.결론 본연구통과대결핵분지간균RV3871기인진행분자극륭,특이단백표체화기인서렬측정,제공료해기인적결구공능특정,병통과생물신식학여동원성대비분석,발현Rv3871기인재치병화비치병분지간균독력분비작용중적결구화공능차이,위결핵병발병궤제천명급기치료약파적사선제공이론화실험의거.
Objective To clone and express the Rv3871 gene related to the virulent protein secretion of Mycobacterium tuberculosis and analyze its molecular structure, function and homology using bioinformatic approach. Methods A pair of primers was designed to amplify the Rv3871 gene, which was subcloned into the prokaryotic plasmid pET32a (+). The recombinant plasmid was identified by sequence analysis and the expressed recombinant protein by SDS-PAGE. The structure, function and homology alignment of Rv3871 were analyzed comparatively against other mycobacteria. Results The restriction fragments through molecular cloning matched perfectly in size with our prediction. The gene sequence was consistent with the corresponding sequence in GenBank. The expression protein was detected by SDS-PAGE with a molecular weight of 84 kD. Two FtsK/SpoE Ⅲ domains were found by bioinformatic analysis. The homology results showed distinct differences between Rv3871 of the pathogenic M. Tuberculosis and its counterparts in non-pathogenic mycobacteria. Conclusions Molecular cloning, expression and sequencing identify the structural and functional characteristics of Rv3871. The structural and functional differences of the gene between pathogenic and non-pathogenic mycobacteria identified by bioinformatics provide some evidence for the pathogenesis and drug targets of tuberculosis.
