Structure of a modular polyketide synthase来源： 时间：六月 19, 2014, 12:33 a.m.
Structure of a modular polyketide synthase
Polyketide natural products constitute a broad class of compounds with diverse structural features and biological activities. Their biosynthetic machinery, represented by type I polyketide synthases (PKSs), has an architecture in which successive modules catalyse two-carbon linear extensions and keto-group processing reactions on intermediates covalently tethered to carrier domains. Here we used electron cryo-microscopy to determine sub-nanometre-resolution three-dimensional reconstructions of a full-length PKS module from the bacterium Streptomyces venezuelae that revealed an unexpectedly different architecture compared to the homologous dimeric mammalian fatty acid synthase. A single reaction chamber provides access to all catalytic sites for the intramodule carrier domain. In contrast, the carrier from the preceding module uses a separate entrance outside the reaction chamber to deliver the upstream polyketide intermediate for subsequent extension and modification. This study reveals for the first time, to our knowledge, the structural basis for both intramodule and intermodule substrate transfer in polyketide synthases, and establishes a new model for molecular dissection of these multifunctional enzyme systems.
Structural rearrangements of a polyketide synthase module during its catalytic cycle
The polyketide synthase (PKS) mega-enzyme assembly line uses a modular architecture to synthesize diverse and bioactive natural products that often constitute the core structures or complete chemical entities for many clinically approved therapeutic agents1. The architecture of a full-length PKS module from the pikromycin pathway ofStreptomyces venezuelae creates a reaction chamber for the intramodule acyl carrier protein (ACP) domain that carries building blocks and intermediates between acyltransferase, ketosynthase and ketoreductase active sites (see accompanying paper2). Here we determine electron cryo-microscopy structures of a full-length pikromycin PKS module in three key biochemical states of its catalytic cycle. Each biochemical state was confirmed by bottom-up liquid chromatography/Fourier transform ion cyclotron resonance mass spectrometry. The ACP domain is differentially and precisely positioned after polyketide chain substrate loading on the active site of the ketosynthase, after extension to the β-keto intermediate, and after β-hydroxy product generation. The structures reveal the ACP dynamics for sequential interactions with catalytic domains within the reaction chamber, and for transferring the elongated and processed polyketide substrate to the next module in the PKS pathway. During the enzymatic cycle the ketoreductase domain undergoes dramatic conformational rearrangements that enable optimal positioning for reductive processing of the ACP-bound polyketide chain elongation intermediate. These findings have crucial implications for the design of functional PKS modules, and for the engineering of pathways to generate pharmacologically relevant molecules.
Structural biology: Enzyme assembly line pictured
Many enzymes form 'assembly lines' containing a series of catalytic modules. Visualization of how the structure of a module shifts during catalysis provides a clearer idea of how such enzymes work.
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