Cloning, expression, and purification of intact polyketide synthase modules.

Autor:	Maschio L; School of Biochemistry, Biomedical Sciences Building, University Walk, University of Bristol, Bristol, United Kingdom; BrisSynBio Synthetic Biology Research Centre, Life Sciences Building, University of Bristol, Bristol, United Kingdom., Parnell AE; School of Biochemistry, Biomedical Sciences Building, University Walk, University of Bristol, Bristol, United Kingdom; BrisSynBio Synthetic Biology Research Centre, Life Sciences Building, University of Bristol, Bristol, United Kingdom., Lees NR; School of Chemistry, Cantock's Close, University of Bristol, Bristol, United Kingdom., Willis CL; School of Chemistry, Cantock's Close, University of Bristol, Bristol, United Kingdom; BrisSynBio Synthetic Biology Research Centre, Life Sciences Building, University of Bristol, Bristol, United Kingdom., Schaffitzel C; School of Biochemistry, Biomedical Sciences Building, University Walk, University of Bristol, Bristol, United Kingdom; BrisSynBio Synthetic Biology Research Centre, Life Sciences Building, University of Bristol, Bristol, United Kingdom., Stach JEM; School of Biology, Ridley Building, Newcastle University, Newcastle, United Kingdom; Centre for Synthetic Biology and the Bioeconomy, Baddiley-Clark Building, Newcastle University, Newcastle, United Kingdom., Race PR; School of Biochemistry, Biomedical Sciences Building, University Walk, University of Bristol, Bristol, United Kingdom; BrisSynBio Synthetic Biology Research Centre, Life Sciences Building, University of Bristol, Bristol, United Kingdom. Electronic address: paul.race@bristol.ac.uk.
Jazyk:	angličtina
Zdroj:	Methods in enzymology [Methods Enzymol] 2019; Vol. 617, pp. 63-82. Date of Electronic Publication: 2019 Feb 04.
DOI:	10.1016/bs.mie.2018.12.018
Abstrakt:	Polyketides are a structurally and functionally diverse family of bioactive natural products that have proven to be a rich source of pharmaceutical and agrochemical lead compounds. Many polyketides are biosynthesized by large multifunctional megaenzymes termed type I modular polyketide synthases (PKSs). These systems possess a distinctive assembly line-like architecture, comprising a series of linearly arranged, multidomain extension modules, housed in sequence within giant polypeptide chains. Due to their inherently modular structures, PKSs represent attractive targets for reengineering, enabling access to functionally optimized "nonnatural" natural products. In this chapter we describe methods for the molecular cloning, recombinant over-expression, and purification of intact PKS modules and multimodular PKS polypeptides. The usefulness of these methods is demonstrated by applying them to the study of the abyssomicin C PKS, a >1MDa multimodular synthase responsible for the biosynthesis of a polyketide antimicrobial lead compound. (© 2019 Elsevier Inc. All rights reserved.)
Databáze:	MEDLINE
Externí odkaz:	Zobrazit plný text záznamu