SPATIAL ARRANGEMENT OF GENE-PRODUCTS OF THE P1 OPERON IN THE MEMBRANEOF MYCOPLASMA-PNEUMONIAE

The spatial arrangement of three P1 operon-encoded proteins-attachment protein P1 (ORF5 gene product) and the ORF6-derived proteins of 40 an d 90 kDa-in the membrane of Mycoplasma pneumoniae M129 was investigate d by nearest-neighbor analysis. For these studies, the homobivalent, t hiol-cleavable, and nonmembrane-permeating cross-linking reagent 3,3'- dithiobis(sulfosuccinimidylpropionate) (DTSSP) was used. The cross-lin ked proteins were isolated by immunoprecipitation with antibodies dire cted against fusion proteins of selected regions of the 40-kDa, the 90 -kDa, or the P1 protein. The individual components of the immunoprecip itated protein complexes were identified by sodium dodecyl sulfate-pol yacrylamide gel electrophoresis, autoradiography, and immunoblot analy sis. This study showed that the P1 protein, the ORF6 gene product, and an unidentified 30-kDa protein were linked to each other in the intac t bacterial membrane by the reagent DTSSP, indicating that these prote ins are located at a maximal distance of 12 A Angstrom (1A Angstrom= 0 .1 nm) on the tip structure of M. pneumoniae.