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Classification of Single Particles from Human Cell Extract Reveals Distinct Structures.


ABSTRACT: Multi-protein complexes are necessary for nearly all cellular processes, and understanding their structure is required for elucidating their function. Current high-resolution strategies in structural biology are effective but lag behind other fields (e.g., genomics and proteomics) due to their reliance on purified samples rather than heterogeneous mixtures. Here, we present a method combining single-particle analysis by electron microscopy with protein identification by mass spectrometry to structurally characterize macromolecular complexes from human cell extract. We identify HSP60 through two-dimensional classification and obtain three-dimensional structures of native proteasomes directly from ab initio classification of a heterogeneous mixture of protein complexes. In addition, we reveal an ?1-MDa-size structure of unknown composition and reference our proteomics data to suggest possible identities. Our study shows the power of using a shotgun approach to electron microscopy (shotgun EM) when coupled with mass spectrometry as a tool to uncover the structures of macromolecular machines.

SUBMITTER: Verbeke EJ 

PROVIDER: S-EPMC6109231 | biostudies-literature | 2018 Jul

REPOSITORIES: biostudies-literature

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Classification of Single Particles from Human Cell Extract Reveals Distinct Structures.

Verbeke Eric J EJ   Mallam Anna L AL   Drew Kevin K   Marcotte Edward M EM   Taylor David W DW  

Cell reports 20180701 1


Multi-protein complexes are necessary for nearly all cellular processes, and understanding their structure is required for elucidating their function. Current high-resolution strategies in structural biology are effective but lag behind other fields (e.g., genomics and proteomics) due to their reliance on purified samples rather than heterogeneous mixtures. Here, we present a method combining single-particle analysis by electron microscopy with protein identification by mass spectrometry to stru  ...[more]

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