Biology subjects

David Matthews

Publications and source records attributed to David Matthews.

3 recordsLinked to original sources

bioRxiv2016

Identification of important amino acid replacements in the 2013-2016 Ebola virus outbreak

The phylogenetic relationships of Zaire ebolavirus have been intensively analysed over the course of the 2013-2016 outbreak. However, there has been limited consideration of the functional impact of this variation. Here we describe an analysis of the available sequence data in the context of protein structure and phylogenetic history. Amino acid replacements are rare and predicted to have minor effects on protein stability. Synonymous mutations greatly outnumber nonsynonymous mutations, and most of the latter fall into unstructured intrinsically disordered regions, indicating that purifying selection is the dominant mode of selective pressure. However, one replacement, occurring early in the outbreak in Gueckedou in Guinea on 31st March 2014 (alanine to valine at position 82 in the GP protein), is close to the site where the virus binds to the host receptor NPC1 and is located in the phylogenetic tree at the origin of the major B lineage of the outbreak. The functional and evolutionary evidence indicates this A82V change likely has consequences for EBOV's host specificity and hence adaptation to humans.

Microbiology↗

bioRxiv2016

Virus genomes reveal the factors that spread and sustained the West African Ebola epidemic.

The 2013-2016 epidemic of Ebola virus disease in West Africa was of unprecedented magnitude, duration and impact. Extensive collaborative sequencing projects have produced a large collection of over 1600 Ebola virus genomes, representing over 5% of known cases, unmatched for any single human epidemic. In a comprehensive analysis of this entire dataset, we reconstruct in detail the history of migration, proliferation and decline of Ebola virus throughout the region. We test the association of geography, climate, administrative boundaries, demography and culture with viral movement among 56 administrative regions. Our results show that during the outbreak viral lineages moved according to a classic gravity model, with more intense migration between larger and more proximate population centers. Despite a strong attenuation of international dispersal after border closures, localized cross-border transmission beforehand had already set the seeds for an international epidemic, rendering these measures relatively ineffective in curbing the epidemic. We use this empirical evidence to address why the epidemic did not spread into neighboring countries, showing that although these regions were susceptible to developing significant outbreaks, they were also at lower risk of viral introductions. Finally, viral genome sequence data uniquely reveals this large epidemic to be a heterogeneous and spatially dissociated collection of transmission clusters of varying size, duration and connectivity. These insights will help inform approaches to intervention in such epidemics in the future.

Epidemiology↗

bioRxiv2016

Allosteric inhibition of Aurora-A kinase by a synthetic VNAR nanobody

The vast majority of clinically-approved protein kinase inhibitors target the ATP binding pocket directly. Consequently, many inhibitors have broad selectivity profiles and most have significant off-target effects. Allosteric inhibitors are generally more selective, but are difficult to identify because allosteric binding sites are often unknown or poorly characterized, and there is no clearly preferred approach to generating hit matter. Aurora-A is activated through binding of TPX2 to an allosteric site on the kinase catalytic domain, and this knowledge could be exploited to generate an inhibitor. However, it is currently unclear how to design such a compound because a small molecule or peptide mimetic of TPX2 would be expected to activate, not inhibit the kinase. Here, we generated an allosteric inhibitor of Aurora-A kinase based on a synthetic, VNAR single domain nanobody scaffold, IgNARV-D01. Biochemical studies and a crystal structure of the Aurora-A/IgNARV-D01 complex show that the nanobody overlaps with the TPX2 binding site. In contrast with the binding of TPX2, which stabilizes an active conformation of the kinase, binding of the nanobody stabilizes an inactive conformation, in which the C-helix is distorted, the canonical Lys-Glu salt bridge is broken, and the regulatory (R-) spine is disrupted by an additional hydrophobic side chain from the activation loop. These studies illustrate how nanobodies can be used to characterize the regulatory mechanisms of kinases and provide a rational basis for structure-guided design of allosteric Aurora-A kinase inhibitors.\n\nSignificanceProtein kinases are commonly dysregulated in cancer and inhibitors of protein kinases are key therapeutic drugs. However, this strategy is often undermined by a lack of selectivity since the ATP binding pocket that kinase inhibitors usually target is highly conserved. Inhibitors that target allosteric sites are more selective but more difficult to generate. Here we identify a single domain antibody (nanobody) to target an allosteric pocket on the catalytic domain of Aurora-A kinase and demonstrate that the mechanism is antagonistic to a physiologically-relevant allosteric activator, TPX2. This work will enable the development of allosteric Aurora-A inhibitors as potential therapeutics, and provide a model for the development of tools to investigate allosteric modes of kinase inhibition.

Biochemistry↗