Biology subjects

Back, C. R.

Publications and source records attributed to Back, C. R..

3 recordsLinked to original sources

bioRxiv2024

Delineation of the Complete Reaction Cycle of a Natural Diels-Alderase

The Diels-Alder reaction is one of the most effective methods for the synthesis of substituted cyclohexenes. The development of protein catalysts for this reaction remains a major priority, affording new sustainable routes to high value target molecules. Whilst a small number of natural enzymes have been shown capable of catalysing [4+2] cycloadditions, there is a need for significant mechanistic understanding of how these prospective Diels-Alderases promote catalysis to underpin their development as biocatalysts for use in synthesis. Here we present a molecular description of the complete reaction cycle of the bona fide natural Diels-Alderase AbyU, which catalyses formation of the spirotetronate skeleton of the antibiotic abyssomicin C. This description is derived from X-ray crystallographic studies of AbyU in complex with a non-transformable synthetic substrate analogue, together with transient kinetic analyses of the AbyU catalysed reaction and computational reaction simulations. These studies reveal the mechanistic intricacies of this enzyme system and establish a foundation for the informed reengineering of AbyU and related biocatalysts.

biochemistry↗

bioRxiv2022

Discovery and biosynthetic assessment of Streptomyces ortus sp nov. isolated from a deep-sea sponge

The deep sea is known to host novel bacteria with the potential to produce a diverse array of undiscovered natural products. Understanding these bacteria is thus of broad interest in ecology and could also underpin applied drug discovery, specifically in the area of antimicrobials. Here, we isolate a new strain of Streptomyces from the tissue of the deep-sea sponge Polymastia corticata collected at a depth of 1869 m from the Gramberg seamount in the Atlantic Ocean. This strain, which was given the initial designation A15ISP2-DRY2T, has a genome size of 9.29 Mb with a GC content of 70.83%. Phylogenomics determined that A15ISP2-DRY2T represents a novel species within the genus Streptomyces as part of the Streptomyces aurantiacus clade. The biosynthetic potential of A15ISP2-DRY2T was assessed relative to other members of the aurantiacus clade via comparative gene cluster family (GCF) analysis. This revealed a clear congruent relationship between phylogeny and GCF content. A15ISP2-DRY2T contains six unique GCFs absent elsewhere in the clade. Culture-based assays were used to demonstrate the antibacterial activity of A15ISP2-DRY2T against two drug-resistant human pathogens. We thus determine A15ISP2-DRY2T to be a novel bacterial species with considerable biosynthetic potential and propose the systematic name Streptomyces ortus sp. nov. Impact StatementThe Streptomyces genus has contributed more to our antibiotic arsenal than any other group of bacteria or fungi. Despite decades of exploration, global analysis has suggested they still possess more undiscovered biosynthetic diversity than any other bacterial group. Isolating novel species of Streptomyces is therefore a priority for antibiotic discovery. Here we isolate a novel strain from a deep-sea sponge and use comparative cluster analysis to identify six biosynthetic clusters unique to our deep-sea strain. This work demonstrates the utility of continuing to isolate novel Streptomyces strains for antibiotic discovery and, for the first time, we used species tree-gene cluster tree reconciliation to assess the contribution of vertical evolution on the biosynthetic gene cluster content of Streptomyces.

genomics↗

bioRxiv2020

Insights into the role of C3d dimers in B cell activation and Staphylococcal immune evasion

Cleavage of C3 to C3a and C3b plays a central role in the generation of complement-mediated defences. Although the thioester-mediated surface deposition of C3b has been well-studied, fluid-phase dimers of C3 fragments remain largely unexplored. Here we present the first X-ray crystal structures of disulphide-linked human C3d dimers and show they undergo structurally-stabilising N-terminal domain swapping when in complex with the Staphylococcus aureus immunomodulator Sbi. Through binding studies and flow cytometric analyses we uncover the physiologically-relevant roles of these dimers in crosslinking complement receptor 2 and modulating B cell activation to potentially promote anergy. This potential induction of cellular tolerance by C3d dimers could contribute to Sbi-mediated S. aureus immune evasion as well as limit autoreactive immune responses under physiological conditions. Thus, insights gained from our findings could inform the design of novel therapies for autoimmune disorders and enhance our understanding surrounding the importance of complement in the fluid phase.

immunology↗