Structure and recognition of acyl carrier protein domains in a modular polyketide synthase

Modular polyketide synthases (PKS) are multidomain enzymes responsible for biosynthesis of polyketides, a medicinally important class of natural products. In the course of polyketide biosynthesis, an acyl carrier protein (ACP) domain anchors the growing polyketide chain and interacts with other catalytic domains as the polyketide undergoes extension and modification. The modular architecture of these PKS enzymes provides an attractive framework for engineering novel synthases. However, not all engineered combinations of PKS domains are productive and impaired domain-domain interactions have been implicated in some of these cases. A better understanding of the nature and specificity of protein-protein interactions between the PKS domains would be valuable for successful design and engineering of novel combinations of domains for efficient production of new biosynthetic products.;To provide a foundation for studying these domain-domain interactions, the solution structure of an ACP domain from a representative modular PKS, 6-deoxyerythronolide B synthase (DEBS), was determined by nuclear magnetic resonance (NMR) spectroscopy. The overall fold of this 10-kD domain consists of a three-helical bundle, with an additional short helix in the second loop contributing to the helical packing of the bundle. The fold is similar to the few available structures of homologous carrier proteins, with some differences in the length and relative orientation of the helices.;Based on the structure of this prototypical ACP domain, homology models were constructed for five other ACP domains of DEBS. Comparison of their steric and electrostatic surfaces led to a proposed model for the recognition of ACP domains based on a pattern of charged residues presented by each ACP at the putative interaction interface centered on helix II. The predictions of this model are consistent with the previously observed specificity of ACP recognition. Further support for the model was obtained from the results of site-directed mutagenesis experiments, which confirmed an important role in ACP recognition for two of the charged residues identified as part of the pattern of putative electrostatic interactions. This approach should allow future studies to quickly define the residues important for domain-domain interactions involving ACP domains, facilitating design of novel productive combinations of PKS domains for engineered polyketide biosynthesis.