Engineering and production of streptokinase in a Bacillus subtilis expression-secretion system.
Streptokinase is one of the major blood-clot-dissolving agents used in many medical treatments. With the cloned streptokinase gene (skc) available, production of the secreted streptokinase from various Bacillus subtilis strains was studied. The use of the six-extracellular-protease-deficient strain, WB600, greatly improved the production yield of the secreted streptokinase. A modified skc which has the original skc promoter and signal sequence replaced with the B. subtilis levansucrase promoter and signal sequence was also constructed. B. subtilis carrying either the wild-type or the modified skc produces streptokinase at a comparable level. Even with WB600 as the expression host, a C-terminally-processed streptokinase was also observed. Through region-specific combinatorial mutagenesis around the C-terminal processing sites, streptokinase derivatives resistant to C-terminal degradation were engineered. One of the derivatives showed a 2.5-fold increase in specific activity and would potentially be a better thrombolytic agent.
Wong
S L
SL
Department of Biological Sciences, University of Calgary, Alberta, Canada.
Ye
R
R
Nathoo
S
S
D000595:Amino Acid Sequence; D001412:Bacillus subtilis / Q000201:enzymology* / Q000235:genetics*; D001483:Base Sequence; D003470:Culture Media / Q000737:chemistry; D004351:Drug Resistance / Q000235:genetics; D010450:Endopeptidases / Q000494:pharmacology; D005818:Genetic Engineering; D005822:Genetic Vectors / Q000235:genetics*; D008969:Molecular Sequence Data; D009154:Mutation / Q000235:genetics; D010957:Plasmids / Q000235:genetics; D013045:Species Specificity; D013300:Streptokinase / Q000378:metabolism*
D003470:Culture Media; D010450:Endopeptidases; D013300:Streptokinase