PARACOCCIDIOIDES-BRASILIENSIS EXPRESSES BOTH GLYCOSYLPHOSPHATIDYLINOSITOL-ANCHORED PROTEINS AND A POTENT PHOSPHOLIPASE-C

This study reports, for the first time, the detection of glycosylphosp hatidylinositol (GPI) membrane anchors in proteins of a pathogenic fun gus, Paracoccidioides brasiliensis. Taking into account that fungal an tigens are found in the sera of paracoccidioidiomycosis patients and t hat cleavage of this glycolipid by phospholipases is a means of select ive protein release, the presence of an enzyme with this property has also been investigated Using a methodological approach in which the pr oteins were immobilized on nitrocellulose, treated with phospholipase C of Trypanosoma brucei and then probed with antibodies which recogniz e the 1,2-cyclic-phosphate inositol moiety formed as a reaction produc t in proteins bearing the glycolipid anchor, it was possible to detect a major glycoprotein in the 80- to 90-kDa range, as well as two other minor species of 66 and 43 kDa. All of them bind to Concanavalin-A an d are also substrates of a very potent fungal phospholipase C which is inhibited by p-chloromercuri-phenylsulfonic acid and is insensitive t o EDTA. The integrity of glycosylphosphatidylinositol anchors in prote ins of P. brasiliensis is impaired by 0.1 M NaOH, a finding indicative of a diacyl glycerolipid moiety which is quite surprising since it is , with the exception of African trypanosomes surface proteins and Torp edo acetylcholinesterase, an uncommon feature among GPIs in general. T he present findings may have implications in the pathology of paracocc idiodomycosis. (C) 1995 Academic Press, Inc.