Isolation and Characterization of Transcription Signal Sequences from Streptococcus thermophilus

Abstract.

Streptococcus thermophilus (ST) chromosomal DNA fragments generated by partial Sau3A digestion were cloned into the unique BamHI site upstream from the promoterless chloramphenicol acetyltransferase (cat) gene of the Escherichia coli (EC) promoter-probe vector pKK520-3. Recombinant plasmids containing ST sequences with transcription-activation activity were isolated from chloramphenicol-resistant (Cm^R) EC transformants. A promoterless Streptomyces antibioticus melanin biosynthesis operon (melC) was inserted immediately downstream from the ST sequence to identify DNA with strong promoter activity. Several ST transcription-activation sequences, termed ST_Ps, were isolated and subcloned, and their nucleic acid sequences determined. The −10 and −35 consensus sequences were identified in these putative ST promoters. Detailed analysis of ST_P3306 sequence data revealed two partial open reading frames (ORFs) with high degrees of homology to prokaryotic GTP-binding protein and DNA repair enzyme, thus providing valuable information for further study on DNA maintenance in this important lactic acid bacterium.