Cloning and characterization of a nonhemolytic phospholipase C gene from Burkholderia pseudomallei.
We cloned and characterized a phosphatidylcholine-hydrolyzing phospholipase C (PC-PLC) gene from Burkholderia pseudomallei. DNA sequence analysis of the gene indicated an open reading frame coding for 700 amino acids with a 34-amino-acid signal peptide. When cleaved, this yields a secreted 73-kDa mature protein. The deduced amino acid sequence exhibited 48% similarity to that of a nonhemolytic PLC from Pseudomonas aeruginosa. The expressed PC-PLC was heat stable, nonhemolytic for sheep erythrocytes, and active between pH 2 and 8. Western blot analysis with sera from melioidosis patients indicated that they produced immunoglobulin M antibodies against this PC-PLC protein.
Korbsrisate
S
S
Departments of Immunology, Faculty of Medicine Siriraj Hospital, Bangkok 10700, Thailand. grsks@mahidol.ac.th
Suwanasai
N
N
Leelaporn
A
A
Ezaki
T
T
Kawamura
Y
Y
Sarasombath
S
S
D000818:Animals; D000907:Antibodies, Bacterial / Q000097:blood; D001483:Base Sequence; D016957:Burkholderia pseudomallei / Q000201:enzymology* / Q000235:genetics* / Q000276:immunology; D003001:Cloning, Molecular; D017931:DNA Primers / Q000235:genetics; D004269:DNA, Bacterial / Q000235:genetics; D005798:Genes, Bacterial; D006461:Hemolysis; D006801:Humans; D007075:Immunoglobulin M / Q000097:blood; D066298:In Vitro Techniques; D008554:Melioidosis / Q000276:immunology / Q000382:microbiology; D008969:Molecular Sequence Data; D015183:Restriction Mapping; D012756:Sheep; D010738:Type C Phospholipases / Q000235:genetics*
D000907:Antibodies, Bacterial; D017931:DNA Primers; D004269:DNA, Bacterial; D007075:Immunoglobulin M; D010738:Type C Phospholipases; C105598:phosphatidylcholine-specific phospholipase C