Abstract: To investigate the role of enolase in the pathogenesis of highly virulent strains from Streptococcus suis serotype 2,its molecular cloning,protein location analysis,enzymatic activity assay,and immune related function were studied. Based on the whole genome sequencing of S. suis 2 05ZYH33,bioinformatics of gene eno encoding enolase was analyzed. The recombinant expression plasmid pET30a∷eno was transformed into Escherichia coli BL21 competent cells,and the expression was induced. Recombinant protein enolase was obtained by the purification with His-Beads and FPLC. Then the enzyme activity of enolase was detected,and its basal metabolic function was identified. Further,FCM was used to analyze the localization of enolase in S. suis 2. Finally,the effect of enolase on the activity of peripheral blood monouclear cells（PBMCs）was detected by MTT test. Homology analysis showed a highly homologous of eno with that in many others. SignalP and TMHMM analysis revealed that enolase had neither signal peptide sequence nor transmembrane domain structure. Molecular cloning and sequencing illustrated that the eno fragment length was 1 308 bp. The 75 kD protein was acquired after the recombinant plasmid was induced to express and purification. Enzyme activity assay showed that the purified recombinant enolase had the ability to convert 2-PEG into PEP. FCM analysis demonstrated that enolase also existed on the surface of bacteria. The results of MTT test showed that enolase resulted in the decrease of PBMCs activity. In conclusion,enolase is not only involved in the activities of glucose metabolism in S. suis 2,but also exists on its surface,probably involving in the infection process by destroying the mononuclear cells.

Key words: Streptococcus suis serotype 2（S. suis 2）, enolase, hemolytic activity, flow cytometry, peripheral blood monouclear cells