The molecular characterization of the first autolytic lysozyme of Streptococcus pneumoniae reveals evolutionary mobile domains

A biochemical approach to identify proteins with high affinity for choline- containing pneumococcal cell walls has allowed the localization, cloning an d sequencing of a gene (lytC) coding for a protein that degrades the cell w alls of Streptococcus pneumoniae. The lytC gene is 1506 bp long and encodes a protein (LytC) of 501 amino acid residues with a predicted M-r of 58682. LytC has a cleavable signal peptide, as demonstrated when the mature prote in (about 55 kDa) was purified from S. pneumoniae. Biochemical analyses of the pure, mature protein proved that LytC is a lysozyme. Combined cell frac tionation and Western blot analysis showed that the unprocessed, primary pr oduct of the lytC gene is located in the pneumococcal cytoplasm whereas the processed, active form of LytC is tightly bound to the cell envelope. In v ivo experiments demonstrated that this lysozyme behaves as a pneumococcal a utolytic enzyme at 30 degrees C. The DNA region encoding the 253 C-terminal amino acid residues of LytC has been cloned and expressed in Escherichia c oli. The truncated protein exhibits a low, but significant, choline-indepen dent lysozyme activity, which suggests that this polypeptide adopts an acti ve conformation. Self-alignment of the N-terminal part of the deduced amino acid sequence of LytC revealed the presence of 11 repeated motifs. These r esults strongly suggest that the lysozyme reported here has changed the gen eral building plan characteristic of the choline-binding proteins of S. pne umoniae and its bacteriophages, i.e. the choline-binding domain and the cat alytic domain are located, respectively, at the N-terminal and the C-termin al moieties of LytC. This work illustrates the natural versatility exhibite d by the pneumococcal genes coding for choline-binding proteins to fuse sep arated catalytic and substrate-binding domains and create new and functiona l mature proteins.