P169: Production of a new octaketide polyene in the calicheamicin biosynthetic pathway

Calicheamicin γ₁^I belongs to a class of enediyne antibiotics that are renowned for their potency and their unusual mechanism of action.  Rearrangement of a bicyclic carbon core produces a diradical species that abstracts two hydrogen atoms from DNA, leading to oxidative damage and double-stranded DNA cleavage.  It is known that a family of highly reducing type I iterative polyketide synthase (PKS) enzymes are responsible for the biosynthetic assembly of the carbon scaffolds that make up the enediyne core structures, but despite recent biochemical advances, the specific details of this construction process remain unknown.

In an effort to understand the early steps of enediyne core biosynthesis, we have focused on the inherent chemical capabilities and selectivities of the calicheamicin enediyne PKS, CalE8.  Heterologous expression of CalE8 in Escherichia coli results in the production of a number of polyketide metabolites, including a series of polyenes that appear to result from premature off-loading from the PKS.  In addition to these products, optimization of fermentation conditions has allowed us to detect and identify a novel octaketide polyene produced by CalE8 that may be relevant to the biosynthesis of calicheamicin.  Moreover, our efforts to characterize PKS-bound products have led to the development of reliable techniques for the chemical release of polyketide intermediates. Key insights into unanticipated environmental conditions that appear to control fundamental aspects of the enediyne PKS biosynthetic pathway will also be discussed.