Nuclear pore complexes (NPCs) mediate nucleocytoplasmic exchange. They are exceptionally large protein complexes that fuse the inner and outer nuclear membranes to form channels across the nuclear envelope. About 30 different protein components, termed nucleoporins, assemble in multiple copies into an intricate cylindrical architecture. Here, we review our current knowledge of the structure of nucleoporins and how those come together in situ. We delineate architectural principles on several hierarchical organization levels, including isoforms, posttranslational modifications, nucleoporins, and higher-order oligomerization of nucleoporin subcomplexes. We discuss how cells exploit this modularity to faithfully assemble NPCs.
Mass spectrometry
(MS) has become one of the key technologies of
structural biology. In this review, the contributions of chemical
cross-linking combined with mass spectrometry (XL-MS) for studying
three-dimensional structures of proteins and for investigating protein–protein
interactions are outlined. We summarize the most important cross-linking
reagents, software tools, and XL-MS workflows and highlight prominent
examples for characterizing proteins, their assemblies, and interaction
networks in vitro and in vivo. Computational
modeling plays a crucial role in deriving 3D-structural information
from XL-MS data. Integrating XL-MS with other techniques of structural
biology, such as cryo-electron microscopy, has been successful in
addressing biological questions that to date could not be answered.
XL-MS is therefore expected to play an increasingly important role
in structural biology in the future.
Highlights d 2-oxo-acid dehydrogenase complexes are studied by integrative analysis of cell extracts d A cryo-EM structure of the active, native pyruvate dehydrogenase complex is resolved d Asymmetric cryo-EM maps reveal enzyme clustering during pyruvate oxidation d A transiently formed catalytic chamber is defined as the ''pyruvate dehydrogenase factory''
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