A Cell-Free Toolkit for Synthetic Biology.

Prokaryotic cell-free coupled transcription-translation (TX-TL) systems are emerging as a powerful tool to examine natural product biosynthetic pathways in a test tube. The key advantages of this approach are the reduced experimental time scales and controlled reaction conditions. To realize this potential, it is essential to develop specialized cell-free systems in organisms enriched for biosynthetic gene clusters.

Modeling the architecture of the regulatory system controlling methylenomycin production in .

Background: The antibiotic methylenomycin A is produced naturally by Streptomyces coelicolor A3(2), a model organism for streptomycetes. This compound is of particular interest to synthetic biologists because all of the associated biosynthetic, regulatory and resistance genes are located on a single cluster on the SCP1 plasmid, making the entire module easily transferable between different bacterial strains. Understanding further the regulation and biosynthesis of the methylenomycin producing gene cluster could assist in the identification of motifs that can be exploited in synthetic regulatory systems for the rational engineering of novel natural products and antibiotics.

Mechanistic insights into class B radical-S-adenosylmethionine methylases: ubiquitous tailoring enzymes in natural product biosynthesis.

Class B radical S-adenosylmethionine (SAM) methylases are notable for their ability to catalyse methylation reactions in the biosynthesis of a wide variety of natural products, including polyketides, ribosomally biosynthesised and post-translationally modified peptides (RiPPs), nonribosomal peptides (NRPs), aminoglycosides, β-lactams, phosphonates, enediynes, aminocoumarins and terpenes. Here, we discuss the diversity of substrates and catalytic mechanism utilised by such enzymes, highlighting the stereochemical course of methylation reactions at un-activated carbon centres and the ability of some members of the family to catalyse multiple methylations.