Characterization of novel, phenol-soluble polypeptides which confer rigidity to the sheath of Methanospirillum hungatei GP1.
Treatment of the Methanospirillum hungatei GP1 sheath with 90% (wt/vol) phenol resulted in the solubilization of a novel phenol-soluble group of polypeptides. These polypeptides were purified by the removal of insoluble material by ultracentrifugation and represented approximately 19% of the mass of the sheath. The phenol-insoluble material resembled untreated sheath but had lost its rigidity and cylindrical form. Recombination of phenol-soluble and phenol-insoluble fractions by dialysis to remove phenol resulted in cylindrical reassembly products. Although bona fide sheath (complete with the 2.8-nm lattice) was not produced, a role for the phenol-soluble polypeptides in the maintenance of sheath rigidity is implied. The phenol-soluble polypeptides have limited surface exposure as detected by antibodies on intact sheath; therefore, they are not responsible for the 2.8-nm repeat occurring on the outer face of the sheath. However, longitudinal and transverse linear labeling by protein A-colloidal gold on the outer and inner faces, respectively, occurred with monoclonal antibodies specific to the phenol-soluble polypeptides. Restricted surface exposure of phenol-soluble polypeptides on the sheath highlighted molecular defects in sheath architecture. These lattice faults may indicate sites of sheath growth to accommodate cell growth or division (longitudinal immunogold label) and filament division (transverse immunogold label). The identification of a second group of polypeptides within the infrastructure of the sheath suggests that the sheath is a trilaminar structure in which phenol-soluble polypeptides are sandwiched between sodium dodecyl sulfate-beta-mercaptoethanol-EDTA-soluble polypeptides (G. Southam and T. J. Beveridge, J. Bacteriol. 173:6213-6222, 1991) (phenol-insoluble material).
Southam
G
G
Department of Microbiology, College of Biological Science, University of Guelph, Ontario, Canada.
Beveridge
T J
TJ
D000911:Antibodies, Monoclonal; D046911:Macromolecular Substances; D008565:Membrane Proteins / Q000737:chemistry* / Q000302:isolation & purification / Q000648:ultrastructure; D017012:Methanobacteriales / Q000737:chemistry* / Q000648:ultrastructure; D016253:Microscopy, Immunoelectron; D010455:Peptides / Q000737:chemistry* / Q000302:isolation & purification; D019800:Phenol; D010636:Phenols; D012995:Solubility; D013329:Structure-Activity Relationship
D000911:Antibodies, Monoclonal; D046911:Macromolecular Substances; D008565:Membrane Proteins; D010455:Peptides; D010636:Phenols; D019800:Phenol