Characterisation of the hydroxystreptomycin phosphotransferase gene (sph) of Streptomyces glaucescens: Nucleotide sequence and promoter analysis

Abstract

The nucleotide sequence of a 1384 bp fragment containing the coding and promoter sequences of the streptomycin phosphotransferase gene (sph) of the hydroxystreptomycin-producing Streptomyces glaucescens was determined. Evidence for an ATG as translation start codon for sph was derived from a comparison with the amino-terminal amino acid sequence of an aminoglycoside phosphotransferase (aphD gene product) of S. griseus, exhibiting a high degree of amino acid homology to the deduced amino acid sequence of the S. glaucescens sph gene product. Transcriptional start and termination sites for the sph gene were identified by primer extension and/or nuclease S1 mapping experiments. The promoter region of the sph gene appears to be complex since tandemly arranged promoters (orfIp1, orfIp2) initiating transcription of a likely coding region (ORFI) in the opposite direction overlap sph promoter sequences. The presumptive sphp and orfIp1 promoters show considerable sequence similarities in the -10 region to Escherichia coli consensus promoter sequences but no homology to E. coli or Streptomyces -35 regions.

Transcriptional Control of the Hydrogen Cyanide Biosynthetic Genes hcnABC by the Anaerobic Regulator ANR and the Quorum-Sensing Regulators LasR and RhlR in Pseudomonas aeruginosa

“…4) was performed by a single-step RNA isolation method (14) using the TRIzol reagent (GIBCO-BRL). After treatment with amplification-grade DNase (Pharmacia), 10-g aliquots of the isolated RNA were analyzed by primer extension as described elsewhere (54). The oligonucleotide P E 5Ј-GAGGTGGAT GGTCATGTCTGCCCGCGAGAG-3Ј (positions ϩ65 to ϩ36), which anneals to the coding strand of hcnA, was 5Ј-end labeled with 20 Ci of [␥-…”

Versatility of Aminoglycosides and Prospects for Their Future

“…Four enzymes that phosphorylate streptomycin at the 6 position have been identified. The genes for two of them, APH(6)-Ia and -Ib, were cloned from streptomycin producers, Streptomyces griseus and Streptomyces glaucescens (72,308). The third enzyme, APH(6)-Ic, is encoded by a gene located within transposon Tn5, which is found infrequently in gram-negative bacteria (268).…”

Nucleotide sequences and heterologous expression of tcmG and tcmP, biosynthetic genes for tetracenomycin C in Streptomyces glaucescens

“…The facts that the cloned tcmP gene complemented the mutation in S. ofivaceus Tu2353R which causes it to produce TCM B3 but no TCM E (51) and that 8-demethyl TCM C is present in S. glaucescens mutants that lack the 8-0-methyltransferase activity (45,67) (67). Biochemical characterization of the three 0-methyltransferases TcmN (57), TcmO (57) (21) and some antibiotic resistance genes (19,40,61). A mutation in the S. coelicolor bidA gene, which is thought to be the source of the cell's only tRNA for the UUA codon (38,40), results in the loss of the ability to form aerial mycelium and to synthesize actinorhodin as well as another S. coelicolor antibiotic (12 (28,30) as well as TCM C (57).…”