novE and novG act as positive regulators of novobiocin biosynthesis.
The biosynthetic gene cluster of the aminocoumarin antibiotic novobiocin contains two putative regulatory genes, i.e., novE and novG. Functional proof for the role of NovG as a positive regulator of novobiocin biosynthesis had been provided previously, and we now investigated the role of novE. Heterologous expression experiments with the novobiocin biosynthetic gene cluster showed that the entire putative promoter region of novE is required to achieve optimal novobiocin production. Overexpression of novE, using a replicative vector, resulted in an increase of novobiocin formation. In contrast, inactivation of novE by in frame deletion resulted in a strong reduction of novobiocin biosynthesis. Novobiocin production could be restored by an intact copy of novE, but also by the regulatory gene novG. These observations suggest that novE is a positive regulator of novobiocin biosynthesis. NovE was expressed in E. coli and purified. However, in contrast to parallel experiments with NovG, no DNA-binding properties could be shown for NovE. RT-PCR experiments showed that expression of novG was detectable in the absence of NovE, and also that expression of novE occurred in absence of NovG.
Dangel
Volker
V
Pharmaceutical Biology, Pharmaceutical Institute, Eberhard-Karls-Universität Tübingen, Auf der Morgenstelle 8, 72076 Tübingen, Germany.
Eustáquio
Alessandra S
AS
Gust
Bertolt
B
Heide
Lutz
L
D000900:Anti-Bacterial Agents / Q000096:biosynthesis*; D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism*; D003001:Cloning, Molecular; D004269:DNA, Bacterial / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000235:genetics / Q000378:metabolism; D004926:Escherichia coli / Q000235:genetics; D017353:Gene Deletion; D018628:Gene Dosage; D015870:Gene Expression; D020869:Gene Expression Profiling; D015964:Gene Expression Regulation, Bacterial; D005816:Genetic Complementation Test; D005810:Multigene Family; D016254:Mutagenesis, Insertional; D009675:Novobiocin / Q000096:biosynthesis*; D011485:Protein Binding; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000302:isolation & purification; D013302:Streptomyces / Q000235:genetics / Q000502:physiology*
D000900:Anti-Bacterial Agents; D001426:Bacterial Proteins; D004269:DNA, Bacterial; D004268:DNA-Binding Proteins; D011994:Recombinant Proteins; D009675:Novobiocin