SARS-CoV-2 has intricate mechanisms for initiating infection, immune evasion/suppression and replication that depend on the structure and dynamics of its constituent proteins. Many protein structures have been solved, but far less is known about their relevant conformational changes. To address this challenge, over a million citizen scientists banded together through the Folding@home distributed computing project to create the first exascale computer and simulate 0.1 seconds of the viral proteome. Our adaptive sampling simulations predict dramatic opening of the apo spike complex, far beyond that seen experimentally, explaining and predicting the existence of 'cryptic' epitopes. Different spike variants modulate the probabilities of open versus closed structures, balancing receptor binding and immune evasion. We also discover dramatic conformational changes across the proteome, which reveal over 50 'cryptic' pockets that expand targeting options for the design of antivirals. All data and models are freely available online, providing a quantitative structural atlas.

BackgroundPichia pastoris has emerged as an important alternative host for producing recombinant biopharmaceuticals, owing to its high cultivation density, low host cell protein burden, and the development of strains with humanized glycosylation. Despite its demonstrated utility, relatively little strain engineering has been performed to improve Pichia, due in part to the limited number and inconsistent frameworks of reported genomes and transcriptomes. Furthermore, the co-mingling of genomic, transcriptomic and fermentation data collected about Komagataella pastoris and Komagataella phaffii, the two strains co-branded as Pichia, has generated confusion about host performance for these genetically distinct species. Generation of comparative high-quality genomes and transcriptomes will enable meaningful comparisons between the organisms, and potentially inform distinct biotechnological utilies for each species.ResultsHere, we present a comprehensive and standardized comparative analysis of the genomic features of the three most commonly used strains comprising the tradename Pichia: K. pastoris wild-type, K. phaffii wild-type, and K. phaffii GS115. We used a combination of long-read (PacBio) and short-read (Illumina) sequencing technologies to achieve over 1000X coverage of each genome. Construction of individual genomes was then performed using as few as seven individual contigs to create gap-free assemblies. We found substantial syntenic rearrangements between the species and characterized a linear plasmid present in K. phaffii. Comparative analyses between K. phaffii genomes enabled the characterization of the mutational landscape of the GS115 strain. We identified and examined 35 non-synonomous coding mutations present in GS115, many of which are likely to impact strain performance. Additionally, we investigated transcriptomic profiles of gene expression for both species during cultivation on various carbon sources. We observed that the most highly transcribed genes in both organisms were consistently highly expressed in all three carbon sources examined. We also observed selective expression of certain genes in each carbon source, including many sequences not previously reported as promoters for expression of heterologous proteins in yeasts.ConclusionsOur studies establish a foundation for understanding critical relationships between genome structure, cultivation conditions and gene expression. The resources we report here will inform and facilitate rational, organism-wide strain engineering for improved utility as a host for protein production.Electronic supplementary materialThe online version of this article (doi:10.1186/s12864-016-2876-y) contains supplementary material, which is available to authorized users.

AbstractThe SARS-CoV-2/COVID-19 pandemic continues to threaten global health and socioeconomic stability. Experiments have revealed snapshots of many of the viral components but remain blind to moving parts of these molecular machines. To capture these essential processes, over a million citizen scientists have banded together through the Folding@home distributed computing project to create the world’s first Exascale computer and simulate protein dynamics. An unprecedented 0.1 seconds of simulation of the viral proteome reveal how the spike complex uses conformational masking to evade an immune response, conformational changes implicated in the function of other viral proteins, and ‘cryptic’ pockets that are absent in experimental snapshots. These structures and mechanistic insights present new targets for the design of therapeutics.This living document will be updated as we perform further analysis and make the data publicly accessible.