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Substrate-assisted catalysis in polyketide reduction proceeds via a phenolate intermediate

DOI: 10.1016/j.chembiol.2016.07.018 DOI Help

Authors: Martin Schafer (John Innes Centre) , Clare E. M. Stevenson (John Innes Centre) , Barrie Wilkinson (John Innes Centre) , David M. Lawson (John Innes Centre) , Mark J. Buttner (John Innes Centre)
Co-authored by industrial partner: No

Type: Journal Paper
Journal: Cell Chemical Biology

State: Published (Approved)
Published: September 2016
Diamond Proposal Number(s): 9475

Open Access Open Access

Abstract: SimC7 is a polyketide ketoreductase involved in biosynthesis of the angucyclinone moiety of the gyrase inhibitor simocyclinone D8 (SD8). SimC7, which belongs to the short-chain dehydrogenase/reductase (SDR) superfamily, catalyzes reduction of the C-7 carbonyl of the angucyclinone, and the resulting hydroxyl is essential for antibiotic activity. SimC7 shares little sequence similarity with characterized ketoreductases, suggesting it might have a distinct mechanism. To investigate this possibility, we determined the structures of SimC7 alone, with NADP+, and with NADP+ and the substrate 7-oxo-SD8. These structures show that SimC7 is distinct from previously characterized polyketide ketoreductases, lacking the conserved catalytic triad, including the active-site tyrosine that acts as central acid-base catalyst in canonical SDR proteins. Taken together with functional analyses of active-site mutants, our data suggest that SimC7 catalyzes a substrate-assisted, two-step reaction for reduction of the C-7 carbonyl group involving intramolecular transfer of a substrate-derived proton to generate a phenolate intermediate.

Journal Keywords: Angucyclines; Simocyclinones; DNA gyrase; DNA topoisomerase; antibiotics; short-chain dehydrogenase/reductase (SDR) superfamily; Ketoreductase; Streptomyces

Diamond Keywords: Enzymes

Subject Areas: Biology and Bio-materials, Chemistry, Medicine

Instruments: I03-Macromolecular Crystallography , I04-1-Macromolecular Crystallography (fixed wavelength) , I04-Macromolecular Crystallography

Other Facilities: No

Added On: 16/09/2016 15:18

Discipline Tags:

Health & Wellbeing Biochemistry Catalysis Chemistry Structural biology Drug Discovery Life Sciences & Biotech

Technical Tags:

Diffraction Macromolecular Crystallography (MX)