DNA sequence sampling of the Streptococcus pneumoniae genome to identify novel targets for antibiotic development.

We initiated a survey of the Streptococcus pneumoniae genome by DNA sequence sampling. More than 9,500 random DNA sequences of approximately 500 bases average length were determined. Partial sequences sufficient to identify approximately 95% of the aminoacyl tRNA synthetase genes and ribosomal protein (rps) genes were found by comparing the database of partial sequences to known sequences from other organisms. Many genes involved in DNA replication, repair, and mutagenesis are present in S. pneumoniae. Genes for the major subunits of RNA polymerase are also present, as are genes for two alternative sigma factors, rpoD and rpoN. Many genes necessary for amino acid or cofactor biosynthesis and aerobic energy metabolism in other bacteria appear to be absent from the S. pneumoniae genome. A number of genes involved in cell wall biosynthesis and septation were identified, including six homologs to different penicillin binding proteins. Interestingly, four genes involved in the addition of D-alanine to lipoteicoic acid in other gram positive bacteria were found, even though the lipoteicoic acid in S. pneumoniae has not been shown to contain D-alanine. The S. pneumoniae genome contains a number of chaperonin genes similar to those found in other bacteria, but apparently does not contain genes involved in the type III secretion commonly observed in gram negative pathogens. The G+C content of S. pneumoniae genomic DNA is approximately 43 mole percent and the size of the genome is approximately 2.0 Mb as determined by pulsed-field gel electrophoresis. Many of the genes identified by sequence sampling have been physically mapped to the 19 different SmaI fragments derived from the S. pneumoniae genome. The database of random genome sequence tags (GSTs) provides the starting material for determining the complete genome sequence, gene disruption analysis, and comparative genomics to identify novel targets for antibiotic development.