The genus Streptomyces harbors a wealth of untapped pharmaceutical
potential within uncharacterized biosynthetic clusters whose transcription may be
silent or their product undetected. Regulation of these clusters is, in part, managed
by global transcription factors, nucleoid-associated proteins, and pathway-specific
regulatory proteins. Consequently, a complicated interplay of regulatory elements
governs secondary metabolite production and a lack of understanding of this has
undermined efforts to screen for novel drug candidates. In an effort to better
understand this regulation, we have focussed on one highly expressed NAP, and one
silent and cryptic biosynthetic cluster. In parallel, we have leveraged our existing
knowledge to improve our screening of Streptomyces for novel bioactive compounds.
Here, we investigate the nucleoid-associated protein Streptomyces integration
host factor (sIHF), which non-specifically binds DNA and impacts development and
antibiotic production. Through targeted mutations, we probe the interfaces of sIHF
thought responsible for its interactions with DNA and assess the relevance of these
interfaces to in vivo activity. Further, we examine the transcriptionally silent
SCO6429-SCO6438 biosynthetic cluster and attempt to activate transcription from
its promoter by manipulating its potential regulatory proteins. Binding sites for the
putative global regulator cAMP-receptor protein (Crp) in close proximity to this
cluster and a potential pathway specific regulator are examined. Finally, we
overexpress Crp in a library of Streptomyces and assay for novel bioactivity against
known clinical pathogens.
Collectively, this work establishes a foundation for further exploring the
regulatory networks within Streptomyces and how they may be influenced to
discover novel pharmaceuticals.ThesisMaster of Science (MSc
