Purification and characterization of lysophospholipase-transacylase of pathogenic fungus Candida albicans.
A lysophospholipase-transacylase was purified to homogeneity from the culture broth of Candida albicans by ammonium sulfate precipitation and chromatographs on DEAE-cellulose, Ultrogel AcA-44, first Mono Q, hydroxyapatite, TSKgel-3000 and second Mono Q columns. The purified protein was a single band (Mr 41,000) as inferred by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. It had a specific activity of 78 mumol/min per mg protein for fatty acid release and 320 mumol/min per mg protein for phosphatidylcholine formation. Fatty acid release obeyed Michaelis-Menten kinetics and the apparent Km was 76 microM of 1-palmitoyl-sn-glycero-3-phosphatidylcholine, but Lineweaver-Burk plots of transacylase activity was parabolic. The ratio of hydrolase to transacylase activity of the purified enzyme was varied depending upon the concentration of lysophosphatidylcholine. Transacylation was prominent at high concentration of substrate and the ratio of hydrolase to transacylase was 0.24. Low concentration of palmitoylcarnitine (50 microM) inhibited markedly phosphatidylcholine formation but stimulated fatty acid release. The degree of esterification of 1-acyllysophosphatidylcholine was altered with mixtures of different molecular species of substrate, demonstrating acyl chain selectivity in the transfer process. These results suggest that C. albicans lysophospholipase-transacylase is different from the corresponding mammalian enzymes in enzymatic properties.
Takahashi
M
M
Department of Biochemistry, Gifu University School of Medicine, Japan.
Banno
Y
Y
Shikano
Y
Y
Mori
S
S
Nozawa
Y
Y
D000217:Acyltransferases / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism*; D000596:Amino Acids / Q000032:analysis; D002176:Candida albicans / Q000201:enzymology*; D006863:Hydrogen-Ion Concentration; D008245:Lysophospholipase / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism*; D009097:Multienzyme Complexes / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism*; D010172:Palmitoylcarnitine / Q000493:pharmacokinetics; D012709:Serum Albumin / Q000493:pharmacokinetics
D000596:Amino Acids; D009097:Multienzyme Complexes; D012709:Serum Albumin; D010172:Palmitoylcarnitine; D000217:Acyltransferases; D008245:Lysophospholipase; C021782:lysophospholipase-transacylase