Transcriptional analysis and heterologous expression of the gene encoding beta-lactamase inhibitor protein (BLIP) from Streptomyces clavuligerus.
Transcription of bli, the gene encoding beta-lactamase (Bla) inhibitor protein (BLIP) of Streptomyces clavuligerus, was analyzed by promoter-probe studies, Northern hybridization and high-resolution S1 nuclease mapping. The 1-kb SalI DNA fragment immediately upstream from the bli open reading frame (ORF) showed promoter activity when tested using the xylE-based promoter-probe vector, pIJ4083. The promoter activity was approx. 36-fold higher in S. clavuligerus than in S. lividans. Northern hybridization analysis of S. clavuligerus RNA revealed that bli was expressed as a 0.7-kb monocistronic transcript. High-resolution S1 nuclease mapping identified the transcription start point as an A residue 47 bp upstream from the bli start codon. When the bli ORF, along with 111 bp of upstream sequence including the promoter, was introduced into S. lividans, the transformants produced BLIP, but in amounts approx. 12-fold lower than that produced by S. clavuligerus. Involvement of some additional regulatory element that is present in S. clavuligerus, but absent in S. lividans, could explain the difference in the promoter activities and therefore the difference in the overall expression of bli in the two hosts.
Paradkar
A S
AS
Department of Microbiology, University of Alberta, Edmonton, Canada.
Petrich
A K
AK
Leskiw
B K
BK
Aidoo
K A
KA
Jensen
S E
SE
D001426:Bacterial Proteins / Q000235:genetics*; D001483:Base Sequence; D015152:Blotting, Northern; D003001:Cloning, Molecular; D004269:DNA, Bacterial; D008969:Molecular Sequence Data; D011401:Promoter Regions, Genetic; D012329:RNA, Bacterial; D013302:Streptomyces / Q000235:genetics* / Q000378:metabolism; D014158:Transcription, Genetic; D065093:beta-Lactamase Inhibitors
D001426:Bacterial Proteins; D004269:DNA, Bacterial; D012329:RNA, Bacterial; D065093:beta-Lactamase Inhibitors; C065596:beta-lactamase-inhibitor protein, Streptomyces