pectate lyase promoting streptolysin o expression in escherichia coli and strengthening its activity

Background: streptolysin o (slo) from streptococcus pyogenes could oligomerize to form large pores on the membrane of some eu- karyotic cells. because of the action, slo had multifaceted applications in animal cell biology and medicine science. in the previous researches, the highest yield of recombinant expressed slo might be no more than 124 mg/l medium. objectives: a fusion protein of pectate lyase and streptolysin o (pel-slo) was highly expressed with the pectate lyase (pel, from aspergillus nidulans) as fusion partner. the expression, purification, and hemolysis characteristics of this fusion protein were re- searched. methods: the slo gene in this study was from s. pyogenes nz131 and the pectate lyase gene from a. nidulans gr5. escherichia coli bl21 (de3) was used as expression host and pet-28a (+) plasmid as expression vector. ni2+ -nitrilotriacetate-agarose column was used for protein purification. the fusion protein activity was measured by monitoring hemolysis of sheep red blood cells. results: in shaking flask fermentation, the goal protein expression might be higher than 600 mg/l culture at 0.6 mm iptg, 25°c, and 200 rpm for 36 hours, about 5 fold of what was previously reported. the purified pel-slo fusion protein had a specialty of 1 × 107 hu/mg, about 10 fold of natural slo. for sheep red blood cell, pel-slo fusion protein exhibited its optimal hemolysis level at ph 6.5 and 30 - 40°c. conclusions: the research demonstrated that pel might have promoted slo expression and strengthened its activity.