Identification and classification of all potential hemolysin encoding genes and their products from Leptospira interrogans serogroup Icterohaemorrhagiae serovar Lai  

作　　者：Yi-xuanZHAN YanGENG BoBI Jian-yongHE Xiao-kuiGUO Guo-pingZHAO 

机构地区：[1]ResearchCenterofBiotechnology,ShanghaiInstitutesforBiologicalSciences,ChineseAcademyofSciences,Shanghai200233,China [3]TheSchoolofPharmaceu-ticalEngineering,ShenyangPharmaceuticalUniversity,Shenyang110016,China [4]DepartmentofMicrobiologyandParasitologyShanghaiSecondMedicalUniversityShanghai200025,China [5]ChineseNationalHumanGenomeCenteratShanghai,ZhangJiangHighTechPark,Shanghai201203,China

出　　处：《Acta Pharmacologica Sinica》2005年第4期453-461,共9页中国药理学报（英文版）

基　　金：Project supported by the National Natural Science Foundation of China (No 30370071);the National High Technoogy Research and Development Program of China(863 Program,No_ 2003,AA223030).

摘　　要：Aim:To identify and classify all potential hemolysin candidates of Leptospira interrogans serogroup Icterohaemorrhagiae serovar Lai.Methods:All of the potential hemolysin encoding genes were characterized in silico.These genes were cloned and expressed in Escherichia coli.The hemolytic activities of the expressed proteins were assayed observing the hemolysis on sheep blood agar plates.Sphingomyelinase activities of the hemolysin candidates were measured by thin-layer chromatography(TLC)and HPLC for sphingomyelin-hydrolysis. Expression and secretion of the hemolysins in L interrogans were studied by reverse transcription polymerase chain reaction,Western blot,and enzyme-linked immunosorbent assays.Results and Conclusion:The hemolytic activities of hemolysin candidates(LA0327,LA0378,LA1027,LA1029,LAI650,LA3050, LA3937,LA4004)from L interrogans strain Lai were confirmed.They were further divided into two groups,sphingomyelinase hemolysins and non-sphingomyelinase hemolysins,based on their ability to hydrolyze sphingomyelin.Most of these hemolysins were actually expressed in living L interrogans and some of them were secreted into the environment.This study establishes an essential find complete basis for further studying the contribution of hemolysins to the patho- genesis of L interrogans.Aim:To identify and classify all potential hemolysin candidates of Leptospira interrogans serogroup Icterohaemorrhagiae serovar Lai.Methods:All of the potential hemolysin encoding genes were characterized in silico.These genes were cloned and expressed in Escherichia coli.The hemolytic activities of the expressed proteins were assayed observing the hemolysis on sheep blood agar plates.Sphingomyelinase activities of the hemolysin candidates were measured by thin-layer chromatography(TLC)and HPLC for sphingomyelin-hydrolysis. Expression and secretion of the hemolysins in L interrogans were studied by reverse transcription polymerase chain reaction,Western blot,and enzyme-linked immunosorbent assays.Results and Conclusion:The hemolytic activities of hemolysin candidates(LA0327,LA0378,LA1027,LA1029,LAI650,LA3050, LA3937,LA4004)from L interrogans strain Lai were confirmed.They were further divided into two groups,sphingomyelinase hemolysins and non-sphingomyelinase hemolysins,based on their ability to hydrolyze sphingomyelin.Most of these hemolysins were actually expressed in living L interrogans and some of them were secreted into the environment.This study establishes an essential find complete basis for further studying the contribution of hemolysins to the patho- genesis of L interrogans.

关 键 词：HEMOLYSIN Leptospira interrogans sphingo myelinase 

分 类 号：R575[医药卫生—消化系统]