Site-directed mutagenesis of the Pseudomonas aeruginosa type III secretion system protein PscJ reveals an essential role for surface-localized residues in needle complex function.
The Pseudomonas aeruginosa type III secretion system (TTSS) protein PscJ belongs to the PrgK family of TTSS proteins. These proteins are predicted to form one of the inner membrane localized ring substructures of the TTSS needle complex. To determine which amino acid residues of PscJ are important for its function, the pscJ gene was subjected to site-directed mutagenesis. Fifteen individual PscJ amino acid residues that are located in conserved regions of the PrgK family were targeted for mutagenesis. Eight of these residues could be subjected to non-conservative substitution mutagenesis without affecting the function of the resultant mutant protein. Substitution of the other 7 residues (E26, K52, E105, A107, G126, H133, and V189) resulted in either a non-functional protein or the loss of detectable protein. When the essential residues were mapped on to the crystal structure of the E. coli PrgK homolog EscJ, the majority appeared to localize to surface-exposed regions of the protein suggesting a role for these regions in the assembly of the PscJ ring structure.
Burns
Robyn E
RE
Department of Microbiology and Molecular Genetics, Oklahoma State University, 307 Life Sciences East, Stillwater, OK 74078, USA.
McDaniel-Craig
Ashley
A
Sukhan
Anand
A
D000595:Amino Acid Sequence; D019943:Amino Acid Substitution; D001426:Bacterial Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D008565:Membrane Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D008958:Models, Molecular; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D021381:Protein Transport; D011550:Pseudomonas aeruginosa / Q000737:chemistry* / Q000235:genetics* / Q000378:metabolism
D001426:Bacterial Proteins; D008565:Membrane Proteins