Properties of an acid phosphatase from Legionella micdadei which blocks superoxide anion production by human neutrophils.
The high-speed supernatant (100,000 g, 1 h) obtained after centrifuging a suspension of Legionella micdadei that had been freeze-thawed and sonicated contained (i) considerable acid phosphatase activity when assayed using 4-methylumbelliferyl phosphate (MUP) as the substrate, and a factor that blocked superoxide anion production by human neutrophils stimulated with f-Met-Leu-Phe. Chromatography of the extract on a hydroxylapatite column resolved two acids phosphatases (designated ACP1 and ACP2). Subsequent chromatography of ACP2 on a Sephadex G-150 column revealed coincident elution of phosphatase activity and neutrophil blocking activity. When heated at 45 degrees C for various periods of time, the phosphatase activity of the acid phosphatase preparation was lost at the same rate as the ability of the preparation to block superoxide anion production by neutrophils. Furthermore, preincubation of neutrophils and acid phosphatase together in the presence of a heteropolymolybdate complex that inhibits the phosphatase eliminated the effect of the L. micdadei phosphatase on neutrophil superoxide anion production. ACP2 had the following properties: pH optimum, 6.0; Km for MUP, 3.8 mM; isoelectric point, 4.5; substrate specificity, MUP greater than ADP greater than phosphoenolpyruvate greater than phosphothreonine greater than phosphoserine greater than phosphotyrosine; molecular weight (estimated by sucrose density gradient centrifugation and gel filtration chromatography), 71,000-86,000. These results indicate that a cell-associated phosphatase may play a role in the virulence of L. micdadei.
D000135:Acid Phosphatase / Q000378:metabolism*; D002850:Chromatography, Gel; D004305:Dose-Response Relationship, Drug; D005615:Freezing; D006358:Hot Temperature; D006801:Humans; D007525:Isoelectric Focusing; D007527:Isoenzymes / Q000378:metabolism; D007700:Kinetics; D007875:Legionella / Q000201:enzymology*; D009504:Neutrophils / Q000378:metabolism*; D013010:Sonication; D013481:Superoxides / Q000096:biosynthesis*
D007527:Isoenzymes; D013481:Superoxides; D000135:Acid Phosphatase