S35: Rational Control of the Direct Products of an Enediyne PKS

Enediyne natural products such as calicheamicin γ₁^I are among the most potent antibiotics known. It has been established that the carbon backbone of the calicheamicin aglycone is synthesized in nature by an iterative polyketide synthase (PKS), yet the precise structure of the direct polyketide precursor of the aglycone skeleton has yet to be identified. Unlike modular PKSs, in which the polyketide product can often be predicted from the primary sequence of proteins involved, iterative PKSs produce advanced polyketides using a single set of biosynthetic domains "programmed" in a precise yet not well-understood way. In general, the biochemical challenge of accessing and characterizing iterative PKS intermediates often confounds efforts to understand the mechanistic details of the natural product's early biosynthetic steps.

Our efforts to characterize PKS-bound products of the calicheamicin enediyne PKS CalE8 have led to the development of reliable techniques for the chemical release of polyketide intermediates. Analysis of the collection of CalE8-bound products by mass spectrometry has revealed a number of key insights, including a surprising role for environmental conditions in modulating the inherent chemistry of the enediyne PKS. This knowledge allows us to experimentally exert control over the product distribution of this multifunctional enzyme and may facilitate the detection and characterization of other proteins involved in the early steps of calicheamicin biosynthesis.