title: Functional domain analysis of the Mycoplasma pneumoniae co-chaperone TopJ.
abstract: Colonization of conducting airways of humans by the prokaryote Mycoplasma pneumoniae is mediated by a differentiated terminal organelle important in cytadherence, gliding motility and cell division. TopJ is a predicted J-domain co-chaperone also having domains unique to mycoplasma terminal organelle proteins and is essential for terminal organelle function, as well as stabilization of protein P24, which is required for normal initiation of terminal organelle formation. J-domains activate the ATPase of DnaK chaperones, facilitating peptide binding and proper protein folding. We performed mutational analysis of the predicted J-domain, central acidic and proline-rich (APR) domain, and C-terminal domain of TopJ and assessed the phenotypic consequences when introduced into an M. pneumoniae topJ mutant. A TopJ derivative with amino acid substitutions in the canonical J-domain histidine-proline-aspartic acid motif restored P24 levels but not normal motility, morphology or cytadherence, consistent with a J-domain co-chaperone function. In contrast, TopJ derivatives having APR or C-terminal domain deletions were less stable and failed to restore P24, but resulted in normal morphology, intermediate gliding motility and cytadherence levels exceeding that of wild-type cells. Results from immunofluorescence microscopy suggest that both the APR and C-terminal domains, but not the histidine-proline-aspartic acid motif, are critical for TopJ localization to the terminal organelle.
authors.affiliation: Department of Microbiology, University of Georgia, Athens, GA 30602, USA.
authors.forename: Jason M
authors.initials: JM
authors.lastname: Cloward
authors.forename: Duncan C
authors.initials: DC
authors.lastname: Krause
mesh_terms: D000595:Amino Acid Sequence; D019943:Amino Acid Substitution / Q000235:genetics; D001422:Bacterial Adhesion; D001426:Bacterial Proteins / Q000235:genetics* / Q000378:metabolism*; D004252:DNA Mutational Analysis; D006801:Humans; D008124:Locomotion; D046529:Microscopy, Electron, Transmission; D008856:Microscopy, Fluorescence; D018832:Molecular Chaperones / Q000235:genetics* / Q000378:metabolism*; D008969:Molecular Sequence Data; D050505:Mutant Proteins / Q000235:genetics / Q000378:metabolism; D009177:Mycoplasma pneumoniae / Q000166:cytology / Q000201:enzymology* / Q000472:pathogenicity / Q000502:physiology*; D015388:Organelles / Q000737:chemistry; D017434:Protein Structure, Tertiary; D017384:Sequence Deletion
chemical_list: D001426:Bacterial Proteins; D018832:Molecular Chaperones; D050505:Mutant Proteins