球形孢子丝菌亲环素B的功能和定位研究  

A study on function and localization of Sporothrix globosa Cyclophilin B

作　　者：梁梓贤 胡玲 吴榕 夏婷婷[1,2] 李美荣[1] 胡旭初[3,4] 黄怀球[1] LIANG Zixian;HU Ling;WU Rong;XIA Tingting;LI Meirong;HU Xuchu;HUANG Huaiqiu(Department of Dermatology and Venereology,the Third Affiliated Hospital of Sun Yat-sen University,Guangzhou,Guangdong 510630,China;Center for Reproductive Medicine,the Third Affiliated Hospital of Sun Yat-sen University,Guangzhou,Guangdong 510630,China;Department of Parasitology,Zhongshan School of Medicine,Sun Yat-sen University,Guangzhou,Guangdong 510080,China;Key Laboratory of Tropical Disease Control,Sun Yat-sen University,Ministry of Education,Guangzhou,Guangdong 510080,China)

机构地区：[1]中山大学附属第三医院皮肤性病科,广东广州510630 [2]中山大学附属第三医院生殖医学中心,广东广州510630 [3]中山大学中山医学院寄生虫教研室,广东广州510080 [4]中山大学热带病防治研究教育部重点实验室,广东广州510080

出　　处：《热带医学杂志》2023年第4期447-452,I0002,F0004,共8页Journal of Tropical Medicine

基　　金：国家自然科学基金(81974300)。

摘　　要：目的研究球形孢子丝菌亲环素B(CypB)的抗原性、免疫原性以及亚细胞定位。方法通过生物信息学方法从球形孢子丝菌基因组数据库中鉴定出CypB基因(Sg.CypB)及其蛋白(Sg.CypB)。通过原核表达的方法表达出重组Sg.CypB(r Sg.CypB)。将纯化的重组蛋白免疫SD大鼠获得抗r Sg.CypB抗体,用ELISA法检测获得的抗体滴度。用获得的抗体对球形孢子丝菌行免疫荧光染色,观察Sg.CypB在球形孢子丝菌上的分布。结果Sg.CypB基因长930 bp,编码的Sg.CypB含有309个氨基酸。预测Sg.CypB具有分泌信号肽和C端跨膜序列,蛋白主体部位位于细胞外,具有典型的肽基脯氨酸顺反异构酶(PPIase)活性,且能检测出原核表达的r Sg.CypB具有PPIase活性。利用大鼠产生的抗r Sg.CypB抗体(滴度达到1∶51200,P<0.05),免疫荧光检测到Sg.CypB位于球形孢子丝菌表面。结论Sg.CypB是位于球形孢子丝菌表面的具有PPIase活性的抗原分子,可能在真菌-宿主相互作用中发挥作用。Objective To study the antigenicity,immunogenicity,and subcellular localization of Cyclophilin B(CypB)protein of Sporothrix globosa.Methods Bioinformatics methods were employed to identify the CypB gene of S.globosa(Sg.CypB gene)and its encoded protein(Sg.CypB protein)from the S.globosa genome database.The recombinant Sg.CypB protein(r Sg.CypB)was expressed by a prokaryotic expression system.Antibodies against Sg.CypB were generated from rats,which were injected with r Sg.CypB.Titers of antibodies were evaluated with ELISA.Immunofluorescence staining of S.globosa was performed with antibodies to examine the subcellular localization of Sg.CypB.Results The Sg.CypB gene was930 bp long,encoding a 309-amino-acid protein(Sg.CypB).It was predicted that Sg.CypB may be a membrane-associated protein with a typical signal peptide and a single transmembrane region at its C-terminus.The majority of Sg.CypB was exposed to an extracellular environment,processing a typical peptidyl-prolyl cis-trans isomerase(PPIase).The PPIase activity could also be detected in r Sg.CypB.With antibodies detection(titer reach 1∶51200,P<0.05),immunofluorescence staining demonstrated Sg.CypB localizing on the S.globosa cell surface.Conclusion Sg.CypB was an antigen with PPIase activity located on the S.globosa surface,making it possible to play an important role in fungus-host cell interaction.

关 键 词：球形孢子丝菌 亲环素B PPIASE 重组蛋白 

分 类 号：R379.9[医药卫生—病原生物学]