Biology subjects

Mühlberg, L.

Publications and source records attributed to Mühlberg, L..

3 recordsLinked to original sources

bioRxiv2023

Structural Host-Virus Interactome Profiling of Intact Infected Cells

Virus-host protein-protein interactions (PPIs) are fundamental to viral infections, yet high-resolution identification within the native context of intact infected cells has remained an unsolved challenge. Here, we developed structural host-virus interactome profiling (SHVIP) that combines in situ cross-linking mass spectrometry with the enrichment of newly synthesized viral proteins from infected cells. We established SHVIP using herpes simplex virus type 1 and obtained 739 PPIs based on 6,194 cross-links from productively infected cells. SHVIP captures PPIs across intracellular compartments and at the intact host endomembrane system. It resolves PPIs to the protein domain level and seamlessly integrates with AlphaFold-based structural modeling, facilitating detailed predictions of PPI sites within structured and intrinsically disordered regions. We show that SHVIP captures parts of the virus-host PPI space that are elusive to traditional interaction proteomics approaches. By selectively disrupting several newly identified virus-host PPIs, we confirm SHVIPs ability to uncover genuine virus-host PPIs in the intact complex environment of infected cells.

microbiology↗

bioRxiv2023

Enhancing Inter-link Coverage in Cross-Linking Mass Spectrometry through Context-Sensitive Subgrouping and Decoy Fusion

In cross-linking mass spectrometry, sensitivity and specificity in assigning mass spectra to cross-links between different proteins (inter-links) remains challenging. Here, we report on limitations of commonly used concatenated target-decoy searches and propose a target-decoy competition strategy on a fused database as a solution. Further, we capitalize on context-divergent error rates by implementing a novel context-sensitive subgrouping strategy. This approach increases inter-link coverage by [~] 30 - 75 % across XL-MS datasets, maintains low error rates, and preserves structural accuracy.

biochemistry↗

bioRxiv2022

Spatial, Quantitative and Functional Deconstruction of Virus and Host Protein Interactions Inside Intact Cytomegalovirus Particles

Herpesviruses assemble large enveloped particles that are difficult to characterize structurally due to their size, fragility and complex proteome with partially amorphous nature. Here we use cross-linking mass spectrometry and quantitative proteomics to derive a spatially resolved interactome map of intact human cytomegalovirus virions. This enabled the de novo allocation of 32 viral proteins into four spatially resolved virion layers, each organized by a dominant viral scaffold protein. The viral protein UL32 engages with all layers in an N-to-C-terminal radial orientation bridging nucleocapsid to viral membrane. In addition, we observed the layer-specific recruitment of 82 host proteins, a subset of which are constitutively and selectively incorporated via specific host-virus interactions. We uncover how the recruitment of PP1 phosphatase and 14-3-3 proteins by UL32 affects early and late steps during viral biogenesis. Collectively, this study provides global structural insights into the native configuration of virus and host protein interactions inside herpesvirus particles.

microbiology↗