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The structure of a polyketide synthase bimodule core.

ABSTRACT: Polyketide synthases (PKSs) are predominantly microbial biosynthetic enzymes. They assemble highly potent bioactive natural products from simple carboxylic acid precursors. The most versatile families of PKSs are organized as assembly lines of functional modules. Each module performs one round of precursor extension and optional modification, followed by directed transfer of the intermediate to the next module. While enzymatic domains and even modules of PKSs are well understood, the higher-order modular architecture of PKS assembly lines remains elusive. Here, we visualize a PKS bimodule core using cryo-electron microscopy and resolve a two-dimensional meshwork of the bimodule core formed by homotypic interactions between modules. The sheet-like organization provides the framework for efficient substrate transfer and for sequestration of trans-acting enzymes required for polyketide production.

SUBMITTER: Tittes YU

PROVIDER: S-EPMC9491710 | biostudies-literature | 2022 Sep

REPOSITORIES: biostudies-literature

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The structure of a polyketide synthase bimodule core.

Tittes Yves U YU Herbst Dominik A DA Martin Solène F X SFX Munoz-Hernandez Hugo H Jakob Roman P RP Maier Timm T

Science advances 20220921 38

Polyketide synthases (PKSs) are predominantly microbial biosynthetic enzymes. They assemble highly potent bioactive natural products from simple carboxylic acid precursors. The most versatile families of PKSs are organized as assembly lines of functional modules. Each module performs one round of precursor extension and optional modification, followed by directed transfer of the intermediate to the next module. While enzymatic domains and even modules of PKSs are well understood, the higher-orde ...[more]

PMID: 36129979

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Project description:Mycobacterium tuberculosis is currently the leading cause of death by any bacterial infection1. The mycolic acid layer of the cell wall is essential for viability and virulence, and the enzymes responsible for its synthesis are therefore front line targets for antimycobacterial drug development2,3. Polyketide synthase 13 (Pks13) is a module comprised of a closely symmetric parallel dimer of chains, each encoding several enzymatic and transport functions, that carries out the condensation of two different very long chain fatty acids to produce mycolic acids that are essential components of the mycobacterial cell wall. Consequently individual enzymatic domains of Pks13 are targets for antimycobacterial drug development4. To understand this machinery, we sought to determine the structure and domain trajectories of the dimeric multi-enzyme Pks13, a 2×198,426 Dalton complex, from protein purified endogenously from mycobacteria under normal growth conditions, to capture it with normal substrates bound trapped 'in action'. Structures of the multi-domain assembly revealed by cryogenic electron microscopy (cryoEM) define the ketosynthase (KS), linker, and acyltransferase (AT) domains, each at atomic resolution (1.8Å), with bound substrates defined at 2.4Å and 2.9Å resolution. Image classification reveals two distinct structures with alternate locations of the N-terminal acyl carrier protein (termed ACP1a, ACP1b) seen at 3.6Å and 4.6Å resolution respectively. These two structures suggest plausible intermediate states, related by a ~60Å movement of ACP1, on the pathway for substrate delivery from the fatty acyl-ACP ligase (FadD32) to the ketosynthase domain. The linking sequence between ACP1 and the KS includes an 11 amino acid sequence with 6 negatively charged side chains that lies in different positively charged grooves on the KS in ACP1a versus ACP1b structures. This charge complementarity between the extended chain and the grooves suggests some stabilization of these two distinct orientations. Other domains are visible at lower resolution and indicate flexibility relative to the KS-AT core. The chemical structures of three bound endogenous long chain fatty acid substrates with their proximal regions defined in the structures were determined by electrospray ionization mass spectrometry. The domain proximities were probed by chemical cross-linking and identified by mass spectrometry. These were incorporated into integrative structure modeling to define multiple domain configurations that transport the very long fatty acid chains throughout the multistep Pks13 mediated synthetic pathway.

Dataset Information

The structure of a polyketide synthase bimodule core.

Publications

The structure of a polyketide synthase bimodule core.

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