Early Steps of the Biosynthesis of the Anticancer Antibiotic Pleurotin.

Pleurotin is a meroterpenoid specialized metabolite made by the fungus , and it is a lead anticancer molecule due to its irreversible inhibition of the thioredoxin-thioredoxin reductase system. Total synthesis of pleurotin has been achieved, including through a stereoselective route; however, its biosynthesis has not been characterized. In this study, we used isotope-labeled precursor feeding to show that the nonterpenoid quinone ring of pleurotin and its congeners is derived from phenylalanine.

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.

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.

Engineering Transcriptional Regulator Effector Specificity Using Computational Design and In Vitro Rapid Prototyping: Developing a Vanillin Sensor.

The pursuit of circuits and metabolic pathways of increasing complexity and robustness in synthetic biology will require engineering new regulatory tools. Feedback control based on relevant molecules, including toxic intermediates and environmental signals, would enable genetic circuits to react appropriately to changing conditions. In this work, variants of qacR, a tetR family repressor, were generated by computational protein design and screened in a cell-free transcription-translation (TX-TL) system for responsiveness to a new targeted effector.

Design and implementation of a biomolecular concentration tracker.

As a field, synthetic biology strives to engineer increasingly complex artificial systems in living cells. Active feedback in closed loop systems offers a dynamic and adaptive way to ensure constant relative activity independent of intrinsic and extrinsic noise. In this work, we use synthetic protein scaffolds as a modular and tunable mechanism for concentration tracking through negative feedback.