Roles of the carboxy-terminal half of Pseudomonas aeruginosa major outer membrane protein OprF in cell shape, growth in low-osmolarity medium, and peptidoglycan association.
OprF, the major outer membrane protein of Pseudomonas aeruginosa, is multifunctional in that it can act as a nonspecific porin, plays a role in the maintenance of cell shape, and is required for growth in a low-osmolarity environment. The latter two structural roles of OprF, and OprF's association with the peptidoglycan, have been proposed to be localized in the carboxy terminus of the protein, based on this region's similarity to members of the OmpA family of proteins. To determine if this is correct, we constructed a series of C-terminally truncated OprF derivatives and examined their effects on P. aeruginosa cell length and growth in low-osmolarity medium. While the C terminus of OprF was required for wild-type cell length and growth in low-osmolarity medium, expression of the N terminus (first 163 amino acids [aa]) also influenced these phenotypes (compared with OprF deficiency). The first 154 to 164 aa of OprF seemed required for stable protein expression, consistent with the existence of a beta-barrel domain in the N terminus of OprF. Greater than 215 aa of the protein were required for strong peptidoglycan association, confirming that residues in the C-terminal end of OprF are required for peptidoglycan binding. OprF deficiency did not affect the in vivo growth of an OprF-deficient strain in a mouse chamber model. Collectively, these data suggest that the C terminus of OprF plays a role in cell length, growth of P. aeruginosa in low-osmolarity media (but not in vivo), and peptidoglycan association, while the N terminus has an influence on the first two characteristics and is additionally important for stable protein expression.
Rawling
E G
EG
Department of Microbiology and Immunology, University of British Columbia, Vancouver, British Columbia, Canada V6T 1W5.
Brinkman
F S
FS
Hancock
R E
RE
D003470:Culture Media; D017353:Gene Deletion; D006358:Hot Temperature; D008623:Mercaptoethanol / Q000378:metabolism; D009994:Osmolar Concentration; D010457:Peptidoglycan / Q000378:metabolism*; D018272:Porins / Q000235:genetics / Q000378:metabolism*; D011550:Pseudomonas aeruginosa / Q000737:chemistry* / Q000254:growth & development / Q000378:metabolism
D003470:Culture Media; D010457:Peptidoglycan; D018272:Porins; D008623:Mercaptoethanol