Identification of the Mature Sequence of the PhrA Signaling Peptide of Streptococcus pneunomiae

            Experiments in Streptococcus pneumoniae (SP) have demonstrated that the phrA/tprA cassette codes for a signaling system that resembles quorum-sensing systems found in other Gram-positive bacteria. The phrA/tprA system is modeled after Phr peptides in the Bacillus species, leading to the prediction that the PhrA protein is secreted and cleaved to release - from its C-terminal domain - a mature 5-7-residue peptide. The mature PhrA peptide is predicted to signal cells by being transported by an oligopeptide permease into the cytoplasm where the peptide can bind to its TprA receptor protein.  TprA is a negative regulator of phrA transcription, and overexpression of PhrA results in increased expression of phrA, suggesting that mature PhrA peptide inhibits TprA activity. The sequence of the mature PhrA peptide has not been elucidated. In order to determine this sequence deletion constructs of phrA have been created that remove C-terminal residues from PhrA, residues predicted to encode the mature peptide. The truncation alleles of phrA will be transformed into a SP strain that contains a phrA-lacZ reporter and is deleted for the wild-type copy of phrA. Expression of the phrA-lacZ in cells containing either full-length or a truncated phrA allele will be measured. It is expected that cells expressing a truncated phrA allele that removes the portion encoding the mature PhrA peptide will have decreased phrA-lacZ. These studies will aid in understanding the mechanism by which PhrA and TprA control gene expression and the role of these proteins in mediating a quorum-sensing signal.