FUNCTIONAL-ANALYSIS OF THE 2-GENE LYSIS SYSTEM OF THE PNEUMOCOCCAL PHAGE CP-1 IN HOMOLOGOUS AND HETEROLOGOUS HOST-CELLS

The two lysis genes cph1 and cpl1 of the streptococcus pneumoniae bact eriophage Cp-1 coding for holin and lysozyme, respectively, have been cloned and expressed in Escherichia coli. Synthesis of the Cph1 holin resulted in bacterial cell death but not lysis. The cph1 gene was able to complement a lambda Sam mutation in the nonsuppressing E. coli HB1 01 strain to produce phage progeny, suggesting that the holins encoded by both phage genes have analogous functions and that the pneumococca l holin induces a nonspecific lesion in the cytoplasmic membrane. Conc omitant expression of both holin and lysin of Cp-1 in E. coli resulted in cell lysis, apparently due to the ability of the Cpl1 lysozyme to hydrolyze the peptidoglycan layer of this bacterium. The functional an alysis of the cph1 and cpl1 genes cloned in a pneumococcal mutant with a complete deletion of the lytA gene, which codes for the S. pneumoni ae main autolysin, provided the first direct evidence that, in this gr am-positive-bacterium system, the Cpl1 endolysin is released to its mu rein substrate through the activity of the Cph1 holin. Demonstration o f holin function was achieved by proving the release of pneumolysin to the periplasmic fraction, which strongly suggested that the holin pro duces a lesion in the pneumococcal membrane.