RESUMO
Cell-free extracts were prepared from Pseudomonas pseudomallei cells by freezing-thawing, sonication, and differential ultracentrifugation. The extracts were subjected to column chromatography with DEAE-sepharose to obtain glycoprotein fractions. The fractions showed acid phosphatase activity to p-nitrophenyl phosphate, tyrosine phosphate, serine phosphate, but not to threonine phosphate. They were highly antigenic when tested by immunofluorescence assay with the sera of melioidosis patients.
Assuntos
Animais , Burkholderia pseudomallei/enzimologia , Sistema Livre de Células , Fracionamento Químico , Cromatografia em Agarose , Humanos , Proteínas Tirosina Fosfatases/isolamento & purificação , UltracentrifugaçãoRESUMO
The liquid culture of Pseudomonas pseudomallei shows a complex feature in in the pH-activity pattern of acid phosphatase, not a single peak curve. There was an evident tendency that the higher activity shifted to the higher pH range with the growth of culture. The culture in the presence of tunicamycin (20 micrograms/ml) showed a decreased activity selectively in the higher pH range, while the activity in the lower pH was more heat-labile. The bacterial cells grown on agar plates containing tunicamycin were more heat-labile than the untreated control cells. The glucosidase-treatment reduced the enzymatic activity (of the phosphatase-active fractions from the living cells) with the shift of the optimum pH to lower pH. These observations together with some collateral findings suggest that the pH-activity pattern of acid phosphatase in P. pseudomallei is associated with the development of precursor enzyme proteins to mature glycoproteins.