Isolation of Pneumocystis carinii gp120 by fibronectin affinity: evidence for manganese dependence.
Pneumocystis carinii is a major opportunistic lung pathogen and a leading cause of death among patients with the human immunodeficiency virus. Adherence of P. carinii to type I alveolar epithelial cells is essential for growth and replication and has been shown to be mediated in part by fibronectin (Fn). To better understand the mechanisms underlying this attachment, P. carinii-Fn interaction was characterized with respect to divalent and monovalent ion concentration and pH using an 125I-Fn binding assay to P. carinii. The results suggest that P. carinii has a receptor for Fn that was partially dependent on Ca2+, enhanced by Mn2+, and diminished somewhat by Mg2+. Additional data demonstrated that P. carinii-Fn interaction was sensitive to ionic strength. The pH profile revealed that P. carinii-Fn interaction increased with decreasing pH. The results from the binding assay provided the rationale for a simple isolation of the Fn receptor from P. carinii using a Fn-affinity column involving nondenaturing conditions. The isolated receptor appeared highly purified by SDS-PAGE analysis, with apparent molecular weights of 114 to 118 kD and 66 kD. Western blot analysis indicated that this receptor was gp120, a major surface glycoprotein of P. carinii. Furthermore, the isolated receptor inhibited Fn binding to P. carinii. Finally, a monoclonal antibody raised against the affinity-purified gp120 blocked Fn binding to P. carinii.
Wisniowski
P
P
Indiana University School of Medicine, Department of Internal Medicine, Indianapolis.
Pasula
R
R
Martin
W J
WJ
D000818:Animals; D000908:Antibodies, Fungal; D000911:Antibodies, Monoclonal; D000918:Antibody Specificity; D002413:Cations, Divalent / Q000494:pharmacology; D002712:Chlorides / Q000494:pharmacology; D005353:Fibronectins / Q000378:metabolism*; D005656:Fungal Proteins / Q000737:chemistry / Q000276:immunology / Q000302:isolation & purification* / Q000378:metabolism; D006801:Humans; D006863:Hydrogen-Ion Concentration; D008345:Manganese / Q000494:pharmacology / Q000502:physiology*; D008970:Molecular Weight; D011010:Pneumocystis / Q000737:chemistry*; D011485:Protein Binding / Q000187:drug effects; D051381:Rats; D017469:Receptors, Fibronectin / Q000737:chemistry / Q000276:immunology / Q000302:isolation & purification* / Q000378:metabolism
D000908:Antibodies, Fungal; D000911:Antibodies, Monoclonal; D002413:Cations, Divalent; D002712:Chlorides; D005353:Fibronectins; D005656:Fungal Proteins; D017469:Receptors, Fibronectin; D008345:Manganese