Identification and classification of all potential hemolysin encoding genes and their products from Leptospira interrogans serogroup Icterohae-morrhagiae serovar Lai.
OBJECTIVE
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.


CONCLUSIONS
The hemolytic activities of hemolysin candidates (LA0327, LA0378, LA1027, LA1029, LA1650, 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 and complete basis for further studying the contribution of hemolysins to the pathogenesis of L interrogans.
Zhang
Yi-xuan
YX
Research Center of Biotechnology, Shanghai Institutes for Biological Sciences, Chinese Academy of Sciences, Shanghai 200233, China.
Geng
Yan
Y
Bi
Bo
B
He
Jian-yong
JY
Wu
Chun-fu
CF
Guo
Xiao-kui
XK
Zhao
Guo-ping
GP
D000595:Amino Acid Sequence; D000818:Animals; D001426:Bacterial Proteins / Q000235:genetics* / Q000378:metabolism; D004926:Escherichia coli / Q000378:metabolism; D015964:Gene Expression Regulation, Bacterial; D005798:Genes, Bacterial; D006460:Hemolysin Proteins / Q000235:genetics* / Q000378:metabolism; D007921:Leptospira interrogans / Q000235:genetics*; D008969:Molecular Sequence Data; D010802:Phylogeny; D017433:Protein Structure, Secondary; D012756:Sheep; D013108:Sphingomyelin Phosphodiesterase / Q000378:metabolism
D001426:Bacterial Proteins; D006460:Hemolysin Proteins; D013108:Sphingomyelin Phosphodiesterase