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Protein Science (2007), 16:2093-2107. Published by Cold Spring Harbor Laboratory Press. Copyright © 2007 The Protein Society
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Solution structure and proposed domain–domain recognition interface of an acyl carrier protein domain from a modular polyketide synthase

Viktor Y. Alekseyev1, Corey W. Liu2, David E. Cane3, Joseph D. Puglisi2,4, and Chaitan Khosla1,5,6

1 Department of Chemistry, Stanford University, Stanford, California 94305, USA
2 Stanford Magnetic Resonance Laboratory, Stanford University School of Medicine, Stanford, California 94305, USA
3 Department of Chemistry, Brown University, Providence, Rhode Island 02912, USA
4 Department of Structural Biology, Stanford University School of Medicine, Stanford, California 94305, USA
5 Department of Chemical Engineering, Stanford University, Stanford, California 94305, USA
6 Department of Biochemistry, Stanford University, Stanford, California 94305, USA

(RECEIVED May 21, 2007; FINAL REVISION July 4, 2007; ACCEPTED July 5, 2007)

Polyketides are a medicinally important class of natural products. The architecture of modular polyketide synthases (PKSs), composed of multiple covalently linked domains grouped into modules, provides an attractive framework for engineering novel polyketide-producing assemblies. However, impaired domain–domain interactions can compromise the efficiency of engineered polyketide biosynthesis. To facilitate the study of these domain–domain interactions, we have used nuclear magnetic resonance (NMR) spectroscopy to determine the first solution structure of an acyl carrier protein (ACP) domain from a modular PKS, 6-deoxyerythronolide B synthase (DEBS). The tertiary fold of this 10-kD domain is a three-helical bundle; an additional short helix in the second loop also contributes to the core helical packing. Superposition of residues 14–94 of the ensemble on the mean structure yields an average atomic RMSD of 0.64 ± 0.09 Å for the backbone atoms (1.21 ± 0.13 Å for all non-hydrogen atoms). The three major helices superimpose with a backbone RMSD of 0.48 ± 0.10 Å (0.99 ± 0.11 Å for non-hydrogen atoms). Based on this solution structure, homology models were constructed for five other DEBS ACP domains. Comparison of their steric and electrostatic surfaces at the putative interaction interface (centered on helix II) suggests a model for protein–protein recognition of ACP domains, consistent with the previously observed specificity. Site-directed mutagenesis experiments indicate that two of the identified residues influence the specificity of ACP recognition.

Keywords: polyketide; acyl carrier protein; NMR; structure; protein interaction; electrostatic; homology modeling; mutagenesis


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