Publications and News

Crimean-Congo Haemorrhagic Fever Virus (CCHFV) is found across Africa, Asia, and the Middle East where it can cause Haemorrhagic outbreaks with high case fatality rates. Central to the viral life cycle is the viral L-protein, a crucial and multifunctional protein which both transcribes and replicates the viral genome. Here, we present the cryoEM structures of an RNA free and a 5′ promoter bound complex, describing the core catalytic RNA-dependent RNA polymerase (RdRp). We observe an RdRp that is substantially larger than related L-proteins and contains domain insertions unique to the nairovirus family. The 5′ RNA promoter is found in a tight RNA hairpin stabilised by a single base pair, with 5′ binding triggering the closure of protein over the RNA. Functional analysis of the endonuclease and RdRp activities reveals an enzyme which is capable of both activities and demonstrate RdRp inhibition by known antiviral nucleosides. These data advance our understanding of the molecular mechanisms behind genome replication and transcription, that will help inform future antiviral development.

Borna disease virus 1 (BoDV-1) is a non-segmented RNA virus with one of the smallest known RNA virus genomes. BoDV-1 replicates in the nucleus of infected cells using a virally encoded polymerase complex composed of the large protein and phosphoprotein. Here, we present the BoDV-1 polymerase complex at resolutions up to 2.8 Å, describing the fully ordered large polymerase protein bound to tetrameric phosphoprotein. The complex is maintained through the ordered C-terminal region of one copy of the phosphoprotein. Analysis of the model reveals a conserved methyltransferase domain, though key S-adenosyl methionine binding residues are missing. While no RNA is observed in our models, analysis of a sample under reaction conditions induces an opening and closing of the template entry and exit channels, respectively. Higher-order polymerase assemblies suggest oligomerisation as a conserved feature of negative strand RNA virus polymerases. We provide a molecular framework to investigate bornavirus replication and transcription.

Polyketide synthases (PKSs) are multidomain enzymatic assembly lines that biosynthesize a wide selection of bioactive natural products from simple building blocks. In contrast to their cis-acyltransferase (AT) counterparts, trans-AT PKSs rely on stand-alone ATs to load extender units onto acyl carrier protein (ACP) domains embedded in the core PKS machinery. Trans-AT PKS gene clusters also encode stand-alone acyl hydrolases (AHs), which are predicted to share the overall fold of ATs but function like type II thioesterases (TE_IIs), hydrolyzing aberrant acyl chains from ACP domains to promote biosynthetic efficiency. How AHs specifically target short acyl chains, in particular acetyl groups, tethered as thioesters to the substrate-shuttling ACP domains, with hydrolytic rather than acyl transfer activity, has remained unclear. To answer these questions, we solved the first structure of an AH and performed structure-guided activity assays on active site variants. Our results offer key insights into chain length control and selection against coenzyme A-tethered substrates, and clarify how the interaction interface between AHs and ACP domains contributes to recognition of cognate and noncognate ACP domains. Combining our experimental findings with molecular dynamics simulations allowed for the construction of a data-driven model of an AH:ACP domain complex. Our results advance the currently incomplete understanding of polyketide biosynthesis by trans-AT PKSs, and provide foundations for future bioengineering efforts to offload biosynthetic intermediates or enhance product yields.