Purification and characterization of secretory phospholipase B, lysophospholipase and lysophospholipase/transacylase from a virulent strain of the pathogenic fungus Cryptococcus neoformans

Abstract

Infection caused by the fungus Cryptococcus neoformans is potentially fatal. A highly active extracellular phospholipase, demonstrating phospholipase B (PLB), lysophospholipase (LPL) and lysophospholipase\transacylase (LPTA) activities, was purified to homogeneity from C. neoformans using (NH % ) # SO % fractionation, and hydrophobic-interaction, anion-exchange and gel-filtration chromatography. All three enzyme activities copurified as a single protein with an apparent molecular mass of 70-90 kDa by SDS\PAGE and 160-180 kDa by gel filtration. The ratio of the three activities remained constant after each purification step. The amino acid composition, as well as the sequences of the N-terminus and of five internal peptide fragments were novel. The protein was an acidic glycoprotein containing Nlinked carbohydrate moieties, with pI values of 5.5 and 3.5. The apparent V max values for PLB and LPL activities were 12.3 and 870 µmol\min per mg of protein respectively ; the corresponding

Recent Progress on Phospholipases: Different Sources, Assay Methods, Industrial Potential and Pathogenicity

“…Evidence implicating phospholipases in host cell penetration, cell lysis and injury by bacteria and fungus have been reported for Rickettsia rickettsii [94], Clostridium perfringens [12], Legionella pneumophila [38][39][40], Helicobacter pylori [55], Yersinia enterocolitica [58], Candida albicans [4], Cryptococcus neoformans [67,68] and A. fumigatus [70].…”

“…Chen et al [67,68,116] examined 50 C. neoformans isolates for extracellular PL activity using an egg yolk-based agar assay. Forty-nine of these isolates produced a pericolonial precipitate indicative of PL activity.…”

Virulence-Associated Enzymes of Cryptococcus neoformans

“…C. neoformans extracellular supernatants contain phospholipase B, phospholipase C, lysophospholipase, and acyltransferase (91)(92)(93), and phospholipase activity supports fungal attachment to host cells (94). Phospholipase B promotes fungal invasion of host tissue (95) and hydrolyzes phospholipids in lung surfactant and the plasma membrane (92,96). Moreover, it contributes to fungal survival by maintaining cell wall integrity (97) and provides nutrients that can be used as sole carbon sources by C. neoformans during the infection (98, 99).…”

Molecular characterization and serodiagnosis analysis of a novel lysophospholipase from Clonorchis sinensis

“…Additionally, it is accepted that phospholipases are generally secreted by bacteria, yeasts, and mammalian animals (Djordjevic et al 2005;Lu et al 2001;Ghannoum 2000;Chen et al 2000). These observations led to a conclusion that the CsLysoPLA protein might be excreted or secreted by adults of C. sinensis.…”