[Biological profiles of recombinant Suilysin].
OBJECTIVE
To provide insights into the role of Suilysin in the pathogenesis of Streptococcus suis by analysing the biological profiles of recombinant Suilysin (rSLY).


METHODS
sly gene of S. suis strain SC22 was cloned into pET30a(+) vector and overexpressed in E. coli Rosseta, and rSLY was purified with Ni2+ affinity chromatography, anion exchange chromatography and gel filtration. The hemolytic unit of rSLY was measured with human red blood cells. The cellular toxicity of rSLY to human peripheral white blood cell and cells derived from fetal heart (HEH), liver (HEL), kidney (HEK), lung (HPL) was detected by lactate dehydrogenase (LDH) release assay. The blocking effects of water-soluble cholesterol, serum and specific antibody on the hemolytic activity of rSLY were detected. The amount of proinflammatory cytokines in the serum of mice stimulated by rSLY was measured by Luminex-100.


RESULTS
The hemolytic unit of rSLY was 0.125 nmol/L, but the cellular toxicity of 1 nmol/L rSLY to human peripheral white blood cell, HEH, HEL, HEK and HPF was 20%-25%. The hemolytic activity of rSLY was blocked completely by equal molar of water-soluble cholesterol. Human serum containing 2.8 mmol/L-5.7 mmol/L of cholesterol in physical condition, could merely blocked 1 nmol/L of rSLY to lyse erythrocytes. The human serum with the addition of rabbit anti-rSLY IgG to a final concentration of 15 mg/ml decreased the hemolysis by 10 nmol/L rSLY from 77% down to 5% successfully, but hemolysis remained 60% when tested with 100 nmol/L rSLY. Although IL-1 beta and TNF-alpha didn't display any observable change, the level of interleukin -6 (IL-6) and KC were increased continually in C57BL/6 mice injected intraperitoneumly with rSLY whereas the IL-6 and KC level of mice in the control group only increased slightly and quickly decreased.


CONCLUSIONS
The data suggest that rSLY could not only lyse cells but also induce strong inflammatory response and damage immune cells recruited to the sites of inflammation. This finding implies that serum could provide light protection against rSLY, and specific antibody show a concentration-dependent protection.
Du
Huamao
H
College of Biotechnology, Southwest University, Chongqing 400715, China. duhamo@swu.edu.cn
Xu
Lingfeng
L
Wang
Xiaoling
X
Li
Xuemei
X
Ye
Changyun
C
Xu
Jianguo
J
D000818:Animals; D002478:Cells, Cultured; D006460:Hemolysin Proteins / Q000235:genetics / Q000378:metabolism* / Q000494:pharmacology; D006801:Humans; D051379:Mice; D008810:Mice, Inbred C57BL; D011817:Rabbits; D051381:Rats; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism* / Q000494:pharmacology; D017011:Streptococcus suis / Q000166:cytology*
D006460:Hemolysin Proteins; D011993:Recombinant Fusion Proteins; C104760:suilysin