TREPONEMA-PALLIDUM RARE OUTER-MEMBRANE PROTEINS - ANALYSIS OF MOBILITY BY FREEZE-FRACTURE ELECTRON-MICROSCOPY

Freeze-fracture and deep-etch electron microscopy were used to investi gate the molecular architecture of the Treponema pallidum outer membra ne (OM). Freeze-fracture electron microscopy of treponemes freshly har vested from rabbit testes revealed that the intramembranous particles (IMPs) in both the concave and convex OM leaflets were distributed int o alternating areas of relatively high and low particle density; in ma ny OM fractures, IMPs formed rows that ran either parallel to or obliq uely across the fracture faces. Statistical analysis (runs test) confi rmed that the IMPs were nonrandomly distributed in both OM leaflets. E xamination of deep-etched specimens revealed that the particles observ ed in freeze-fractured OMs also were surface exposed. Combined analysi s of deep-etched and cross-fractured treponemes revealed that the OM p articles were located in regions of the OM away. from the endoflagella and closely apposed to the cytoplasmic membrane-peptidoglycan complex . When treponemes were incubated for extended periods with heat-inacti vated immune rabbit syphilitic serum, no alteration in the distributio n of OM IMPs was detected. In further experiments, approximately 1:1 m ixtures of T. pallidum and Escherichia coli or separate suspensions of the nonpathogenic Treponema phagedenis biotype Reiter were fixed at 3 4 degrees C or after cooling to 0 degrees C (to induce lateral phase s eparations that would aggregate IMPs). Only particles in the T. pallid um OM failed to aggregate in cells fixed at the lower temperature. The combined data suggest that the mobility of T. pallidum rare OM protei ns is limited, perhaps as a result of interactions between their perip lasmic domains and components of the peptidoglycan-cytoplasmic membran e complex.