Molecular analysis of the dnaK locus of Leptospira interrogans serovar Copenhageni

• Published in: Gene Vol. 216; no. 1; pp. 21 - 29
• Main Authors: Ballard, Susan A, Go, Margareta, Segers, Ruud P.A.M, Adler, Ben
• Format: Journal Article
• Language: English
• Published: Netherlands Elsevier B.V 17.08.1998
• Subjects: Amino Acid Sequence; Base Sequence; Blotting, Northern; CIRCE; Cloning, Molecular; DNA, Bacterial - chemistry; DNA, Bacterial - genetics; Escherichia coli Proteins; Genes, Bacterial - genetics; Heat shock proteins; HSP70 Heat-Shock Proteins - analysis; HSP70 Heat-Shock Proteins - genetics; Leptospira interrogans - chemistry; Leptospira interrogans - genetics; Molecular Sequence Data; Regulatory Sequences, Nucleic Acid; RNA, Bacterial - analysis; RNA, Bacterial - genetics; Sequence Analysis, DNA; Spirochete; Transcription, Genetic - genetics; aa, amino acid(s); HS, heat shock; nt, nucleotide(s); PAGE, polyacrylamide gel electrophoresis; Spirochete; Heat shock proteins; oligo, oligodeoxyribonucleotide(s); RBS, ribosome binding site(s); RT, reverse transcriptase; CIRCE; tsp, transcription start point(s)
• ISSN: 0378-1119; 1879-0038
• DOI: 10.1016/S0378-1119(98)00329-1

Analysis of the dnaK locus of Leptospira interrogans serovar Copenhageni identified four genes in the order hrcA, grpE, dnaK and dnaJ. This is the first time a homologue of hrcA has been identified in a spirochete. The hrcA gene and a regulatory sequence, designated CIRCE, play a significant role in the regulation of the dnaK locus of several Gram+ organisms. Their presence upstream of dnaK in Leptospira suggested a similiar regulatory mechanism. Transcriptional analysis using reverse transcriptase–PCR demonstrated transcription of all four genes and indicated that hrcA and grpE were co-transcribed, as were grpE and dnaK. Whilst hrcA, grpE and dnaK were closely linked on the chromosome, transcription terminators between dnaK and dnaJ and downstream of dnaJ suggested that this latter gene exists in its own operon. Primer extension analysis located functional promoters upstream of hrcA and grpE; however, no evidence of a functional promoter could be found for dnaJ. Moreover, transcripts encompassing the first three genes or the entire locus could not be demonstrated, suggesting that the four genes are regulated independently at the transcriptional level. These results indicate that the regulation of the dnaK locus of Leptospira differs somewhat from that observed in other organisms.