Association of lipids with integral membrane surface proteins of Mycoplasma hyorhinis.
Triton X-114 (TX-114)-phase fractionation was used to identify and characterize integral membrane surface proteins of the wall-less procaryote Mycoplasma hyorhinis GDL. Phase fractionation of mycoplasmas followed by analysis by sodium dodecyl sulfate-polyacrylamide gel electrophoresis revealed selective partitioning of approximately 30 [35S]methionine-labeled intrinsic membrane proteins into the TX-114 phase. Similar analysis of [3H]palmitate-labeled cells showed that approximately 20 proteins of this organism were associated with lipid, all of which also efficiently partitioned as integral membrane components into the detergent phase. Immunoblotting and immunoprecipitation of TX-114-phase proteins from 125I-surface-labeled cells with four monoclonal antibodies to distinct surface epitopes of M. hyorhinis identified surface proteins p120, p70, p42, and p23 as intrinsic membrane components. Immunoprecipitation of [3H]palmitate-labeled TX-114-phase proteins further established that surface proteins p120, p70, and p23 (a molecule that mediates complement-dependent mycoplasmacidal monoclonal antibody activity) were among the lipid-associated proteins of this organism. Two of these proteins, p120 and p123, were acidic (pI less than or equal to 4.5), as shown by two-dimensional isoelectric focusing. This study established that M. hyorhinis contains an abundance of integral membrane proteins tightly associated with lipids and that many of these proteins are exposed at the external surface of the single limiting plasma membrane. Monoclonal antibodies are reported that will allow detailed analysis of the structure and processing of lipid-associated mycoplasma proteins.
Bricker
T M
TM
Department of Microbiology, School of Medicine, University of Missouri-Columbia 65212.
Boyer
M J
MJ
Keith
J
J
Watson-McKown
R
R
Wise
K S
KS
D000911:Antibodies, Monoclonal; D000942:Antigens, Bacterial / Q000032:analysis*; D000954:Antigens, Surface / Q000032:analysis*; D001426:Bacterial Proteins / Q000032:analysis*; D003902:Detergents; D007457:Iodine Radioisotopes; D007526:Isoelectric Point; D008565:Membrane Proteins / Q000032:analysis*; D008970:Molecular Weight; D009174:Mycoplasma / Q000032:analysis*; D010168:Palmitates / Q000378:metabolism; D012995:Solubility
D000911:Antibodies, Monoclonal; D000942:Antigens, Bacterial; D000954:Antigens, Surface; D001426:Bacterial Proteins; D003902:Detergents; D007457:Iodine Radioisotopes; D008565:Membrane Proteins; D010168:Palmitates