Figure 1. Biosynthesis of 6-deoxyerythronolide B (6dEB) on the DEBS assembly line.

(A) DEBS is comprised of six extension modules (M1-6), a loading didomain (LD), and a terminal thioesterase domain (TE). These enzymes are dispersed among three homodimeric polypeptides (DEBS1-3). Successive polypeptides in the assembly line associate through specific docking domain interactions localized near the N- and C-termini. The LD initiates polyketide synthesis with a propionyl-CoA derived primer that is incrementally elaborated as it traverses M1-M6. The TE releases 6dEB via concomitant cyclization. (B) The chain elongation modules are comprised of homologous domains. Acyl transferase (AT) domains transfer methylmalonyl extender units to their acyl carrier protein (ACP) partner domains. The ACP then associates with the ketosynthase (KS) domain from the same module to enable elongation of the polyketide chain. Following elongation, the ACP-bound chain can be modified by auxiliary domains such as the ketoreductase (KR), dehydratase (DH; not shown), and the enoyl reductase (ER; not shown). The fully processed polyketide intermediate is eventually translocated from this ACP to the KS domain of the downstream module. (C) SDS-PAGE analysis of purified proteins prior to SEC and SAXS. Protein samples are as follows: 1) holo-ACP3, 2) KR1, 3) TE, 4) KSAT3, 5) KSAT3+KR3, 6) M3, 7) M3+TE, and 8) DEBS3. (D) Schematic representation of the constructs analyzed in this study using SAXS. All but one construct in the series is derived from M3. By fusing the TE domain onto M3, the M3+TE homodimer is capable of catalyzing multiple turnover in vitro (18). The bimodular construct DEBS3 is as shown in (A). Domain coloring is consistent where possible throughout the manuscript.