Chiu, W. | bioRxiv Science

bioRxiv2019

Evidence for influenza B virus hemagglutinin adaptation to the human host: high cleavability, acid-stability and preference for cool temperature

Influenza A virus (IAV) and influenza B virus (IBV) cause yearly epidemics with significant morbidity and mortality. When zoonotic IAVs enter the human population, the viral hemagglutinin (HA) requires adaptation to achieve sustained virus transmission. In contrast, IBV has been circulating in humans, its only host, for a long period of time. Whether this entailed adaptation of IBV HA to the human airways is unknown. To address this question, we compared seasonal IAV (A/H1N1 and A/H3N2) and IBV viruses (B/Victoria and B/Yamagata lineage) with regard to host-dependent activity of HA as the mediator of membrane fusion during viral entry. We first investigated proteolytic activation of HA, by covering all type II transmembrane serine protease (TTSP) and kallikrein enzymes, many of which proved present in human respiratory epithelium. Compared to IAV, the IBV HA0 precursor is cleaved by a broader panel of TTSPs and activated with much higher efficiency. Accordingly, knockdown of a single protease, TMPRSS2, was sufficient to abrogate spread of IAV but not IBV in human respiratory epithelial cells. Second, the HA fusion pH proved similar for IBV and human-adapted IAVs (one exception being HA of 1918 IAV). Third, IBV HA exhibited higher expression at 33{degrees}C, a temperature required for membrane fusion by B/Victoria HA. This indicates pronounced adaptation of IBV HA to the mildly acidic pH and cooler temperature of human upper airways. These distinct and intrinsic features of IBV HA are compatible with extensive host-adaptation during prolonged circulation of this respiratory virus in the human population.\n\nImportanceInfluenza epidemics are caused by influenza A (IAV) and influenza B (IBV) viruses. IBV causes substantial disease, however it is far less studied than IAV. While IAV originates from animal reservoirs, IBV circulates in humans only. Virus spread requires that the viral hemagglutinin (HA) is active and sufficiently stable in human airways. We here resolve how these mechanisms differ between IBV and IAV. Whereas human IAVs rely on one particular protease for HA activation, this is not the case for IBV. Superior activation of IBV by several proteases should enhance shedding of infectious particles. IBV HA exhibits acid-stability and a preference for 33{degrees}C, indicating pronounced adaptation to the human upper airways, where the pH is mildly acidic and a cooler temperature exists. These adaptive features are rationalized by the long existence of IBV in humans, and may have broader relevance for understanding the biology and evolution of respiratory viruses.

microbiology↗

bioRxiv2019

Measurement of Atom Resolvability in CryoEM Maps with Q-scores

CryoEM density maps are now at the point where resolvability of individual atoms can be achieved. However, resolvability is not necessarily uniform throughout the map. We introduce a quantitative parameter to characterize the resolvability of individual atoms in cryoEM maps, the map Q-score. Q-scores can be calculated for atoms in proteins, nucleic acids, water, ligands, and other solvent atoms, using models fitted to or derived from cryoEM maps. Q-scores can also be averaged to represent larger features such as entire residues and nucleotides. Averaged over entire models, Q-scores correlate very well with the estimated resolution of cryoEM maps for both protein and RNA. Assuming the models they are calculated from are well-fitted to the map, Q-scores can thus be used as another measure to indicate resolvability of features in cryoEM maps at various scales, from entire complexes down to individual atoms. Q-score analysis of multiple cryoEM maps of the same proteins derived from different labs confirms reproducibility of structural features down to water and ion atoms.

biochemistry↗

bioRxiv2019

Ribosolve: Rapid determination of three-dimensional RNA-only structures

The discovery and design of biologically important RNA molecules is dramatically outpacing three-dimensional structural characterization. To address this challenge, we present Ribosolve, a hybrid method integrating moderate-resolution cryo-EM maps, chemical mapping, and Rosetta computational modeling, and demonstrate its application to thirteen previously unknown 119-to 338-nucleotide protein-free RNA-only structures: full-length Tetrahymena ribozyme, hc16 ligase with and without substrate, full-length V. cholerae and F. nucleatum glycine riboswitch aptamers with and without glycine, Mycobacterium SAM-IV riboswitch with and without S-adenosylmethionine, and computer-designed spinach-TTR-3, eterna3D-JR_1, and ATP-TTR-3 with and without AMP. Blind challenges, prospective compensatory mutagenesis, internal controls, and simulation benchmarks validate the Ribosolve models and establish that modeling convergence is quantitatively predictive of model accuracy. These results demonstrate that RNA-only 3D structure determination can be rapid and routine.

biophysics↗