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Title: A nanobody suite for yeast scaffold nucleoporins provides details of the nuclear pore complex structure

Abstract

Nuclear pore complexes (NPCs) are the main conduits for molecular exchange across the nuclear envelope. The NPC is a modular assembly of ~500 individual proteins, called nucleoporins or nups. Most scaffolding nups are organized in two multimeric subcomplexes, the Nup84 or Y complex and the Nic96 or inner ring complex. Working in S. cerevisiae, and to study the assembly of these two essential subcomplexes, we here develop a set of twelve nanobodies that recognize seven constituent nucleoporins of the Y and Nic96 complexes. These nanobodies all bind specifically and with high affinity. We present structures of several nup-nanobody complexes, revealing their binding sites. Additionally, constitutive expression of the nanobody suite in S. cerevisiae detect accessible and obstructed surfaces of the Y complex and Nic96 within the NPC. Overall, this suite of nanobodies provides a unique and versatile toolkit for the study of the NPC.

Authors:
ORCiD logo [1]; ORCiD logo [1];  [1];  [2]; ORCiD logo [2]; ORCiD logo [1]
  1. Massachusetts Inst. of Technology (MIT), Cambridge, MA (United States)
  2. Harvard Medical School, Boston, MA (United States)
Publication Date:
Research Org.:
Argonne National Lab. (ANL), Argonne, IL (United States). Advanced Photon Source (APS)
Sponsoring Org.:
USDOE Office of Science (SC), Basic Energy Sciences (BES); National Institutes of Health (NIH)
OSTI Identifier:
1736296
Grant/Contract Number:  
AC02-06CH11357; R01GM77537; T32GM007287; GM103403
Resource Type:
Accepted Manuscript
Journal Name:
Nature Communications
Additional Journal Information:
Journal Volume: 11; Journal Issue: 1; Journal ID: ISSN 2041-1723
Publisher:
Nature Publishing Group
Country of Publication:
United States
Language:
ENGLISH
Subject:
59 BASIC BIOLOGICAL SCIENCES; nuclear pore complex; protein design; proteins; x-ray crystallography

Citation Formats

Nordeen, Sarah A., Andersen, Kasper R., Knockenhauer, Kevin E., Ingram, Jessica R., Ploegh, Hidde L., and Schwartz, Thomas U.. A nanobody suite for yeast scaffold nucleoporins provides details of the nuclear pore complex structure. United States: N. p., 2020. Web. https://doi.org/10.1038/s41467-020-19884-6.
Nordeen, Sarah A., Andersen, Kasper R., Knockenhauer, Kevin E., Ingram, Jessica R., Ploegh, Hidde L., & Schwartz, Thomas U.. A nanobody suite for yeast scaffold nucleoporins provides details of the nuclear pore complex structure. United States. https://doi.org/10.1038/s41467-020-19884-6
Nordeen, Sarah A., Andersen, Kasper R., Knockenhauer, Kevin E., Ingram, Jessica R., Ploegh, Hidde L., and Schwartz, Thomas U.. Wed . "A nanobody suite for yeast scaffold nucleoporins provides details of the nuclear pore complex structure". United States. https://doi.org/10.1038/s41467-020-19884-6. https://www.osti.gov/servlets/purl/1736296.
@article{osti_1736296,
title = {A nanobody suite for yeast scaffold nucleoporins provides details of the nuclear pore complex structure},
author = {Nordeen, Sarah A. and Andersen, Kasper R. and Knockenhauer, Kevin E. and Ingram, Jessica R. and Ploegh, Hidde L. and Schwartz, Thomas U.},
abstractNote = {Nuclear pore complexes (NPCs) are the main conduits for molecular exchange across the nuclear envelope. The NPC is a modular assembly of ~500 individual proteins, called nucleoporins or nups. Most scaffolding nups are organized in two multimeric subcomplexes, the Nup84 or Y complex and the Nic96 or inner ring complex. Working in S. cerevisiae, and to study the assembly of these two essential subcomplexes, we here develop a set of twelve nanobodies that recognize seven constituent nucleoporins of the Y and Nic96 complexes. These nanobodies all bind specifically and with high affinity. We present structures of several nup-nanobody complexes, revealing their binding sites. Additionally, constitutive expression of the nanobody suite in S. cerevisiae detect accessible and obstructed surfaces of the Y complex and Nic96 within the NPC. Overall, this suite of nanobodies provides a unique and versatile toolkit for the study of the NPC.},
doi = {10.1038/s41467-020-19884-6},
journal = {Nature Communications},
number = 1,
volume = 11,
place = {United States},
year = {2020},
month = {12}
}

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