Structure of the native Sec61 protein-conducting channel

Pfeffer, Stefan; Burbaum, Laura; Unverdorben, Pia; Pech, Markus; Chen, Yuxiang; Zimmermann, Richard; Beckmann, Roland; Förster, Friedrich

doi:10.1038/ncomms9403

Cited by 176 publications

(193 citation statements)

References 36 publications

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“…Recent cryo-electron tomographic studies showed the relative positions of mammalian translocons and OSTs bound to the ribosomes 24,26 . However, the detailed interface—i.e., which subunits mediate the OST/translocon interaction—was unclear.…”

Section: Introductionmentioning

confidence: 99%

The atomic structure of a eukaryotic oligosaccharyltransferase complex

Bai

Wang

Zhao

et al. 2018

Nature

101

131

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SUMMARY N-glycosylation is a ubiquitous modification of eukaryotic secretory and membrane-bound proteins; about 90% of glycoproteins are N-glycosylated. The reaction is catalyzed by an eight-protein oligosaccharyltransferase complex, OST, embedded in the ER membrane. Our understanding of eukaryotic protein N-glycosylation has been limited due to the lack of high-resolution structures. Here we report a 3.5-Å resolution cryo-EM structure of the Saccharomyces cerevisiae OST, revealing the structures of Ost1–5, Stt3, Wbp1, and Swp1. We found that seven phospholipids mediate many of the inter-subunit interactions, and an Stt3 N-glycan mediates interaction with Wbp1 and Swp1 in the lumen. Ost3 was found to mediate the OST-Sec61 translocon interface, funneling the acceptor peptide towards the OST catalytic site as the nascent peptide emerges from the translocon. The structure provides novel insights into co-translational protein N-glycosylation and may facilitate the development of small-molecule inhibitors targeting this process.

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Section: Introductionmentioning

confidence: 99%

The atomic structure of a eukaryotic oligosaccharyltransferase complex

Bai

Wang

Zhao

et al. 2018

Nature

101

131

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“…2B). Notably, cryo-tomography of the Sec61 complex in native ER microsomes shows a similar configuration [16]. Although the heterogeneous and uncertain functional state of the cryotomography structure complicates its placement within the translocation cycle [17], it does illustrate that our engaged structure is likely to be compatible with a membrane environment.…”

mentioning

confidence: 96%

Structure of the Sec61 channel opened by a signal sequence

Voorhees

Hegde

2016

Science

207

301

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Secreted and integral membrane proteins comprise up to one-third of the biological proteome. These proteins contain hydrophobic signals that direct their translocation across or insertion into the lipid bilayer by the Sec61 protein conducting channel. The molecular basis for how hydrophobic signals within a nascent polypeptide trigger channel opening is not understood. Here, we use electron cryo-microscopy to determine the structure of an active Sec61 channel that has been opened by a signal sequence. The signal supplants helix 2 of Sec61α, triggering a rotation that opens the central pore both axially across the membrane and laterally toward the lipid bilayer.Comparisons to structures of Sec61 in other states suggest a pathway for how hydrophobic signals engage the channel to gain access to the lipid bilayer.The universally conserved Sec complex forms a gated protein translocation channel at the eukaryotic endoplasmic reticulum (ER) and bacterial plasma membrane [1]. The central component of this channel, SecY in bacteria and Sec61α in eukaryotes, contains ten transmembrane (TM) helices arranged around a central pore [2]. Two single-TM subunits in eukaryotes, Sec61β and Sec61γ, are peripheral to Sec61α. The central pore in the inactive Sec61α/SecY is occluded by a short "'plug"' helix that must be displaced to allow translocation. The interface where TM helices 2/3 contact helices 7/8 defines a "lateral gate" for membrane access of polypeptides [1][2][3].Crystal structures of the Sec complex [2, 4-6] lack a translocating polypeptide and likely represent a range of inactive states. Depending on crystal contacts or translocation partners, the lateral gate and plug are in various states of opening and displacement. However, the biological relevance of these channel conformations has been difficult to interpret without a well-resolved and matched active structure. Previous structures of translocation or insertion intermediates of the ribosome-Sec complex determined by electron cryo-microscopy (cryo-EM) were of moderate resolution [7][8][9], contained heterogeneous substrates [9], required artificial stabilization [8], or were at an uncertain stage of insertion [7]. While these earlier structures were the first views of substrate-induced structural changes consistent with lateral

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“…In eukaryotes, the insertion of most membrane proteins occurs through the ER translocon, which performs its activity assisted by multiple associated proteins. Among them, the TRAP complex is a stoichiometric component of the ribosome-bound translocon complex [44], which has been postulated to participate in the early stages of protein translocation [9], and in moderating topogenic determinants at a final stage of membrane protein biogenesis in vivo [11]. Interestingly, structural data showed that membrane-embedded regions of TRAP complex and Sec61 translocon are in close proximity, which does not block the lateral gate of Sec61 translocon that permits TM segment partition into the lipid bilayer [45].…”

Section: Discussionmentioning

confidence: 99%

Membrane insertion and topology of the translocon-associated protein (TRAP) gamma subunit

Bañó‐Polo

Martínez-Garay

Grau

et al. 2017

Biochimica et Biophysica Acta (BBA) - Biomembranes

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Translocon-associated protein (TRAP) complex is intimately associated with the ER translocon for the insertion or translocation of newly synthesised proteins in eukaryotic cells. The TRAP complex is comprised of three singlespanning and one multiple-spanning subunits. We have investigated the membrane insertion and topology of the multiple-spanning TRAP-γ subunit by glycosylation mapping and green fluorescent protein fusions both in vitro and in cell cultures. Results demonstrate that TRAP-γ has four transmembrane (TM) segments, an Nt/Ct cytosolic orientation and that the less hydrophobic TM segment inserts efficiently into the membrane only in the cellular context of full-length protein.

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Structure of the native Sec61 protein-conducting channel

Cited by 176 publications

References 36 publications

The atomic structure of a eukaryotic oligosaccharyltransferase complex

The atomic structure of a eukaryotic oligosaccharyltransferase complex

Structure of the Sec61 channel opened by a signal sequence

Membrane insertion and topology of the translocon-associated protein (TRAP) gamma subunit

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