Genetic interaction mapping informs integrative structure determination of protein complexes

Braberg, Hannes; Echeverria, Ignacia; Bohn, Stefan; Cimermančič, Peter; Shiver, Anthony L.; Alexander, Richard; Xu, Jiewei; Shales, Michael; Dronamraju, Raghuvar; Jiang, Shuangying; Dwivedi, Gajendradhar; Bogdanoff, Derek; Chaung, Kaitlin; Hüttenhain, Ruth; Wang, Shuyi; Mavor, David; Pellarin, Riccardo; Schneidman, Dina; Bader, Joel S.; Fraser, James S.; Morris, John H.; Haber, James E.; Strahl, Brian D.; Gross, Carol A.; Dai, Junbiao; Boeke, Jef D.; Šali, Andrej; Krogan, Nevan J.

doi:10.1126/science.aaz4910

Cited by 26 publications

(28 citation statements)

References 144 publications

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“…A number of schemes for searching for good-scoring models are available, including local refinement methods and stochastic sampling methods in the continuous space of Cartesian coordinates as well as enumeration schemes in the discrete spaces of both Cartesian and internal coordinates. Finally, IMP implements protocols for analyzing and validating models, including methods for model clustering based on their structural similarity ( 4 , 5 , 9 , 17 , 19 , 29 , 33 , 34 , 36 , 45 , 66 , 68 , 86 , 121 , 122 , 123 , 124 , 125 , 126 , 127 , 128 , 129 , 130 , 131 , 131 , 132 , 132 , 133 , 134 , 135 ). IMP, Integrative Modeling Platform .…”

Section: Software For Integrative Modelingmentioning

confidence: 99%

From integrative structural biology to cell biology

Šali

2021

Journal of Biological Chemistry

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Section: Software For Integrative Modelingmentioning

confidence: 99%

From integrative structural biology to cell biology

Šali

2021

Journal of Biological Chemistry

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“…Since many macromolecular assemblies still remain unresolvable by traditional structural biology techniques, it is necessary to integrate complementary experimental data 42 and theoretical approaches [43][44][45][46][47][48] . In addition, 3D structures often use truncated protein constructs and provide snapshots of the studied complexes outside their cellular environment 28,43 .…”

Section: Discussionmentioning

confidence: 99%

A quantitative mapping approach to identify direct interactions within complexomes

Trepte

Secker

Choi

et al. 2021

Preprint

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Complementary methods are required to fully characterize all protein complexes, or the complexome, of a cell. Affinity purification coupled to mass-spectrometry (AP-MS) can identify the composition of complexes at proteome-scale. However, information on direct contacts between subunits is often lacking. In contrast, solving the 3D structure of protein complexes can provide this information, but structural biology techniques are not yet scalable for systematic, proteome-wide efforts. Here, we optimally combine two orthogonal high-throughput binary interaction assays, LuTHy and N2H, and demonstrate that their quantitative readouts can be used to differentiate direct interactions from indirect associations within multiprotein complexes. We also show that LuTHy allows accurate distance measurements between proteins in live cells and apply these findings to study the impact of the polyglutamine expansion mutation on the structurally unresolved N-terminal domain of Huntingtin. Thus, we present a new framework based on quantitative interaction assays to complement structural biology and AP-MS techniques, which should help to provide first-approximation contact maps of multiprotein complexes at proteome-scale.Graphical Abstract

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“…As the field of XL-MS moves toward intracellular applications, the potential for studying host–pathogen complexes, pathways, and networks on an unbiased level becomes more feasible. Complementary to this in vivo XL-MS application is the use of in vivo genetic interactions obtained using the point mutant epistatic miniarray profile (pE-MAP) platform to derive spatial restraints for integrative modeling ( 110 , 111 , 112 ). In such cases, the structure of host–pathogen complexes and how they affect infection can be studied by introducing specific mutations into the pathogenic genome and studying the phenotypic consequences using genetic interaction profiling of relevant host genes.…”

Section: Discussionmentioning

confidence: 99%

Characterization of an A3G-VifHIV-1-CRL5-CBFβ Structure Using a Cross-linking Mass Spectrometry Pipeline for Integrative Modeling of Host–Pathogen Complexes

Kaake

Echeverria

Kim

et al. 2021

Molecular & Cellular Proteomics

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Structural analysis of host–pathogen protein complexes remains challenging, largely due to their structural heterogeneity. Here, we describe a pipeline for the structural characterization of these complexes using integrative structure modeling based on chemical cross-links and residue–protein contacts inferred from mutagenesis studies. We used this approach on the HIV-1 Vif protein bound to restriction factor APOBEC3G (A3G), the Cullin-5 E3 ring ligase (CRL5), and the cellular transcription factor Core Binding Factor Beta (CBFβ) to determine the structure of the (A3G-Vif-CRL5-CBFβ) complex. Using the MS-cleavable DSSO cross-linker to obtain a set of 132 cross-links within this reconstituted complex along with the atomic structures of the subunits and mutagenesis data, we computed an integrative structure model of the heptameric A3G-Vif-CRL5-CBFβ complex. The structure, which was validated using a series of tests, reveals that A3G is bound to Vif mostly through its N-terminal domain. Moreover, the model ensemble quantifies the dynamic heterogeneity of the A3G C-terminal domain and Cul5 positions. Finally, the model was used to rationalize previous structural, mutagenesis and functional data not used for modeling, including information related to the A3G-bound and unbound structures as well as mapping functional mutations to the A3G-Vif interface. The experimental and computational approach described here is generally applicable to other challenging host–pathogen protein complexes.

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Genetic interaction mapping informs integrative structure determination of protein complexes

Cited by 26 publications

References 144 publications

From integrative structural biology to cell biology

From integrative structural biology to cell biology

A quantitative mapping approach to identify direct interactions within complexomes

Characterization of an A3G-VifHIV-1-CRL5-CBFβ Structure Using a Cross-linking Mass Spectrometry Pipeline for Integrative Modeling of Host–Pathogen Complexes

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