Summary auto-generated
Researchers cloned and sequenced the dnaA gene region of Streptococcus pneumoniae, a pathogenic bacterium. The 6-kilobase fragment contained seven open reading frames (ORFs), including genes homologous to known bacterial replication and cell division genes: htrA (serine protease), spo0J (chromosome partitioning), dnaA (replication initiator), and dnaN (DNA polymerase III subunit). The study identified three clusters of DnaA boxes—regulatory sequences where the DnaA protein binds to initiate DNA replication. Two clusters flanked the dnaA gene while a third was located upstream near the htrA gene. This organization, combined with AT-rich regions characteristic of replication origins, suggests the chromosomal origin of replication is located in this region. The findings were localized on the pneumococcal physical chromosome map. The DnaA protein of S. pneumoniae showed high conservation with DnaA proteins from other bacteria, particularly Bacillus subtilis (38.8% identity). The study also identified previously undescribed genes in pneumococci, including htrA, which may play a role in bacterial virulence.
Key findings
- The dnaA gene region of S. pneumoniae contains seven ORFs organized as tRNAArg-orf1-htrA-spo0J-dnaA-dnaN-orfX-orfY with three clusters of DnaA boxes
- Three DnaA box clusters flank the dnaA gene and an AT-rich region, suggesting this location is the chromosomal replication origin
- S. pneumoniae DnaA protein shares 32.5-38.8% amino acid identity with DnaA from other bacteria and contains conserved domains for ATP binding and DNA binding
- The htrA gene, encoding a serine protease, has not been previously described in S. pneumoniae and may contribute to bacterial virulence
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Abstract
The dnaA gene region of Streptococcus pneumoniae was cloned and sequenced. A tRNA gene, seven ORFs and three DnaA box clusters were identified. The order of the genes and intergene regions found was tRNA(Arg)-orf1-DnaA box cluster 3-htrA-spoOJ-DnaA box cluster 2-dnaA- DnaA box cluster 1-dnaN-orfX-orfY. Five ORFs are homologous to known bacterial genes. The tRNA(Arg) gene and orf1, also called orfL, have already been described in pneumococci and have been reported to be preceded by the competence regulation locus comCDE. In Escherichia coli, htrA encodes a serine protease. In Bacillus subtilis, spoOJ plays a role in sporulation and partition. dnaA encodes an initiator replication protein, very well conserved in several bacteria and dnaN encodes the beta subunit of DNA polymerase III in E. coli. The function of orfX is unknown. The N-terminal part of another reading frame, orfY, revealed high homology with a GTP-binding protein, DnaA box clusters were found upstream and downstream from dnaA. The presence of two such clusters suggests that the chromosomal origin of S. pneumoniae is located within this region. The position of dnaA, and therefore the putative origin of replication, were localized on the physical map of S. pneumoniae.