摘要
目的 分离、克隆人泛素结合酶基因 ,并初步研究其组织表达谱。方法 根据与鼠泛素结合酶有较高一致性的人表达序列标签 (EST)设计筛库引物 ,筛选人胎脑 c DNA文库 ,采用生物信息学方法分析筛选到的阳性克隆 ;设计探针 ,应用人多组织杂交膜分析该基因的组织表达谱。结果 从文库中筛选到两个互为剪接异构体的长度分别为 1471和 1315 bp的 c DNA克隆 ,其开放读码框分别编码 147和 10 9个氨基酸 ,与鼠、果蝇、线虫等生物的泛素结合酶均高度同源 ,含泛素结合结构域 ,Gen Bank登录号分别为 AF12 5 0 44和 AF12 5 0 45。Northern杂交显示 ,该基因在成人心脏、胎盘、胰腺等组织中呈特异表达 ,而在脑、肝、肺、肾和骨胳肌中基本不表达。结论 泛素结合酶基因在人体中的表达受到时间和空间的双重调控。
Objective To identify and clone the gene encoding human ubiquitin binding enzyme and study its expression spectrum. Methods According to the sequence of human EST,which is highly homologous to the mouse ubiquitin conjugating enzyme(E2),primers used for library screening were synthesized to screen the human fetal brain cDNA library.The gene was analyzed by making use of bioinformatics and its expression spectrum was studied by using multiple tissue Northern blot. Results Two cDNA clones encoding human ubiquitin conjugating enzyme were isolated and identified.Both containing the ubiquitin conjugating domain,they were 88% identical in amino acid sequences and found to be isoforms of each other with only an exon excised from the short sequence.They belonged to a highly conserved,and widely expressed E2 enzyme family. Northern blot showed that they were expressed exclusively in heart,placenta,and pancreas while no transcripts could be detected in brain,lung,liver,skeletal muscle,or kidney. Conclusions The gene encoding human ubiquitin binding enzyme is expressed under both temporal and spatial control.As a key enzyme in the degradation of proteins,ubiquitin conjugating enzymes play a central role in the expression regulation on the level of post translation.
出处
《中国医学科学院学报》
CAS
CSCD
北大核心
2000年第4期306-311,共6页
Acta Academiae Medicinae Sinicae
基金
国家自然科学基金!( 3 983 0 0 70 )
国家高技术研究发展计划 863重点项目基金!( Z19-0 2 -0 2 -0 1)
国家重点基础性研究 973项目基
