Cloning and characterization of a phosphate-regulated promoter involved in phosphate control of candicidin biosynthesis.
Phosphate strongly repressed the formation of p-aminobenzoic acid (PABA) synthase, an enzyme involved in candicidin biosynthesis. Expression in Streptomyces lividans of the pabS gene (encoding PABA synthase) of Streptomyces griseus is repressed by phosphate at concentrations above 0.1 mM. However, expression of the pabS gene in Escherichia coli is not regulated by phosphate. Phosphate control of the expression of the pabS gene was observed in all plasmids containing the original 4.5-kb BamHI fragment, whereas no phosphate regulation was found when an upstream 1-kb fragment that carries the pabS promoter was deleted. Using the promoter-probe plasmid pIJ424, a '114-bp' promoter was cloned. Expression of the promoterless kanamycin phosphotransferase gene when fused to the '114-bp' promoter was strongly reduced by phosphate (90% at 5 mM concentration). The '114-bp' promoter has been sequenced and the first transcribed nucleotide identified by S1 mapping. The '114-bp' fragment is A + T-rich (54%), as compared to the Streptomyces genome (70-73% GC). The presence of a phosphate control sequence (pcs) in the upstream region of the pabS gene is proposed.
Rebollo
A
A
Departamento de Ecología, Genética y Microbiología, Facultad de Biología, Universidad de León, Spain.
Gil
J A
JA
Liras
P
P
Asturias
J A
JA
Martín
J F
JF
D000935:Antifungal Agents / Q000096:biosynthesis*; D001483:Base Sequence; D002174:Candicidin / Q000096:biosynthesis*; D003001:Cloning, Molecular; D004269:DNA, Bacterial / Q000302:isolation & purification; D004926:Escherichia coli / Q000235:genetics; D005796:Genes; D019868:Kanamycin Kinase; D008969:Molecular Sequence Data; D010710:Phosphates / Q000502:physiology*; D010770:Phosphotransferases / Q000235:genetics / Q000378:metabolism; D010957:Plasmids; D011401:Promoter Regions, Genetic; D015183:Restriction Mapping; D013305:Streptomyces griseus / Q000235:genetics; D000637:Transaminases / Q000235:genetics; D014158:Transcription, Genetic; D014169:Transformation, Bacterial
D000935:Antifungal Agents; D004269:DNA, Bacterial; D010710:Phosphates; D002174:Candicidin; C022309:PABA synthase; D000637:Transaminases; D010770:Phosphotransferases; D019868:Kanamycin Kinase