Isolation of a 48.5-kDa membrane protein from Pseudomonas maltophilia which exhibits immunologic cross-reaction to the beta-subunit of human chorionic gonadotropin.
In separate studies we have shown that Pseudomonas maltophilia (American Type Culture Collection 13637) possesses an immunoglobulin Fc-binding protein. We have found that this protein prevents the application of immunoassays using monoclonal antibodies to study possible production of a CG-like material by this bacteria. Employing an immunoglobulin saturation technique to block this protein as well as a zwitterion detergent membrane solubilization technique, we now report the isolation of a membrane protein from Pseudomonas maltophilia which shows immunological relationships to the beta-subunit and carboxyl tail of human pregnancy CG. This pseudomonas immunoreactive material produced dose-response curves in the following CG immunoassays: 1) a polyclonal rabbit anti-CG equilibrium assay, 2) carboxyl tail CG equilibrium assay, and 3) two CG equilibrium assays using monoclonal antibodies. The pseudomonas CG-like protein did not react in equilibrium assays for human TSH, human LH, or free alpha-subunit of CG. The pseudomonas CG-like protein was purified by affinity chromatography. The purified protein showed only 0.25% cross-reaction with pregnancy CG in the monoclonal antibody equilibrium assays. Furthermore, the purified protein showed no binding to rat testicular CG/LH receptors, but showed avid binding to the pseudomonas CG-binding protein previously described by Richert and Ryan. The pseudomonas protein showed no binding to Concanavalin-A, which avidly binds pregnancy CG. When assessed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and Western blotting, this protein had a mol wt of 48,500 daltons, which is larger than the mol wt of the unglycosylated beta-subunit of pregnancy CG. We conclude that pseudomonas contain a protein that has partial homology to the beta-subunit of pregnancy CG. This material does not bind to mammalian CG receptors, but does bind to a pseudomonas CG-binding site. The latter suggests it has a function, as yet unknown, in pseudomonas.
Grover
S
S
Department of Internal Medicine, University of Utah School of Medicine, Salt Lake City 84132.
McGee
Z A
ZA
Odell
W D
WD
D001426:Bacterial Proteins / Q000737:chemistry / Q000276:immunology / Q000302:isolation & purification*; D015153:Blotting, Western; D006063:Chorionic Gonadotropin / Q000276:immunology*; D002845:Chromatography; D003429:Cross Reactions; D004591:Electrophoresis, Polyacrylamide Gel; D010450:Endopeptidases / Q000378:metabolism; D006801:Humans; D008565:Membrane Proteins / Q000737:chemistry / Q000276:immunology / Q000302:isolation & purification*; D008970:Molecular Weight; D011549:Pseudomonas / Q000378:metabolism*; D012995:Solubility
D001426:Bacterial Proteins; D006063:Chorionic Gonadotropin; D008565:Membrane Proteins; D010450:Endopeptidases