Production of pneumolysin, a pneumococcal toxin, in Bacillus subtilis.
We have expressed the pneumolysin gene of Streptococcus pneumoniae in Bacillus subtilis, both from its own promoter and as a fusion protein. The level of expression of pneumolysin from its own promoter was low. The protein produced was hemolytically active. A higher level of expression (about 10 micrograms/ml of culture) was achieved when either one of two C-terminal fragments (corresponding to amino acids 265-471 and 55-471, respectively) or the entire coding part of the pneumolysin gene were fused to the promoter and signal sequence-coding region of the alpha-amylase gene of Bacillus amyloliquefaciens. The C-terminal fusion peptides reacted with anti-pneumolysin serum, but were not hemolytically active. In both cases most of the peptide remained cell-associated. When the entire pneumolysin gene was fused to the signal sequence, a hemolytically active form of pneumolysin could be detected, and most of the product was found in a processed form in the culture supernatant. The full-length pneumolysin secreted from B. subtilis was partially purified and used as antigen in an enzyme immunoassay with rabbit anti-pneumolysin serum.
Taira
S
S
National Public Health Institute, Helsinki, Finland.
Jalonen
E
E
Paton
J C
JC
Sarvas
M
M
Runeberg-Nyman
K
K
D001412:Bacillus subtilis / Q000235:genetics*; D001426:Bacterial Proteins; D002850:Chromatography, Gel; D003001:Cloning, Molecular; D004591:Electrophoresis, Polyacrylamide Gel; D004797:Enzyme-Linked Immunosorbent Assay; D005798:Genes, Bacterial; D010957:Plasmids; D011401:Promoter Regions, Genetic; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis / Q000235:genetics; D012689:Sequence Homology, Nucleic Acid; D013296:Streptococcus pneumoniae / Q000235:genetics; D013301:Streptolysins / Q000096:biosynthesis* / Q000235:genetics / Q000302:isolation & purification; D014169:Transformation, Bacterial; D000516:alpha-Amylases / Q000096:biosynthesis / Q000235:genetics
D001426:Bacterial Proteins; D011993:Recombinant Fusion Proteins; D013301:Streptolysins; C032290:plY protein, Streptococcus pneumoniae; D000516:alpha-Amylases