한빛사논문
- 조회1,461
Abstract
Dae In Kima, Birendra KCa, Wenhong Zhub, Khatereh Motamedchabokib, Valerie Doyec, and Kyle J. Rouxa,d,1
aSanford Children's Health Research Center, Sanford Research, Sioux Falls, SD 57104;
bSanford-Burnham Proteomics Facility, Sanford-Burnham Medical Research Institute, La Jolla, CA 92037;
cInstitut Jacques Monod, Unite Mixte de Recherche 7592, Centre National de la Recherche Scientifique, Universite Paris Diderot, Sorbonne Paris Cite, F-75205 Paris, France; and
dDepartment of Pediatrics, Sanford School of Medicine, University of South Dakota, Sioux Falls, SD 57105
Abstract
Proximity-dependent biotin identification (BioID) is a method for identifying protein associations that occur in vivo. By fusing a promiscuous biotin ligase to a protein of interest expressed in living cells, BioID permits the labeling of proximate proteins during a defined labeling period. In this study we used BioID to study the human nuclear pore complex (NPC), one of the largest macromolecular assemblies in eukaryotes. Anchored within the nuclear envelope, NPCs mediate the nucleocytoplasmic trafficking of numerous cellular components. We applied BioID to constituents of the Nup107-160 complex and the Nup93 complex, two conserved NPC subcomplexes. A strikingly different set of NPC constituents was detected depending on the position of these BioID-fusion proteins within the NPC. By applying BioID to several constituents located throughout the extremely stable Nup107?160 subcomplex, we refined our understanding of this highly conserved subcomplex, in part by demonstrating a direct interaction of Nup43 with Nup85. Furthermore, by using the extremely stable Nup107-160 structure as a molecular ruler, we defined the practical labeling radius of BioID. These studies further our understanding of human NPC organization and demonstrate that BioID is a valuable tool for exploring the constituency and organization of large protein assemblies in living cells.
1To whom correspondence should be addressed.
Author contributions: D.I.K., V.D., and K.J.R. designed research; D.I.K., B.K., W.Z., and K.M. performed research; D.I.K., W.Z., K.M., V.D., and K.J.R. analyzed data; and V.D. and K.J.R. wrote the paper.
논문정보
- Research article
- 2014년 06월 (BRIC 등록일 2014-06-10)
- 국외 연구진
댓글 작성 로그인
팝업 닫기관련 링크
