Membrane protein biogenesis faces the challenge of chaperoning hydrophobic transmembrane helices for faithful membrane insertion. The guided entry of tail-anchored proteins (GET) pathway targets and inserts tail-anchored (TA) proteins into the endoplasmic reticulum (ER) membrane with an insertase (yeast Get1/Get2 or mammalian WRB/CAML) that captures the TA from a cytoplasmic chaperone (Get3 or TRC40, respectively). Here, we present cryo-electron microscopy reconstructions, native mass spectrometry, and structure-based mutagenesis of human WRB/CAML/TRC40 and yeast Get1/Get2/Get3 complexes. Get3 binding to the membrane insertase supports heterotetramer formation, and phosphatidylinositol binding at the heterotetramer interface stabilizes the insertase for efficient TA insertion in vivo. We identify a Get2/CAML cytoplasmic helix that forms a "gating" interaction with Get3/TRC40 important for TA insertion. Structural homology with YidC and the ER membrane protein complex (EMC) implicates an evolutionarily conserved insertion mechanism for divergent substrates utilizing a hydrophilic groove. Thus, we provide a detailed structural and mechanistic framework to understand TA membrane insertion.

Structural Basis of Tail-Anchored Membrane Protein Biogenesis by the GET Insertase Complex / Mcdowell, M. A.; Heimes, M.; Fiorentino, F.; Mehmood, S.; Farkas, A.; Coy-Vergara, J.; Wu, D.; Bolla, J. R.; Schmid, V.; Heinze, R.; Wild, K.; Flemming, D.; Pfeffer, S.; Schwappach, B.; Robinson, C. V.; Sinning, I.. - In: MOLECULAR CELL. - ISSN 1097-4164. - 80:1(2020), pp. 72-86. [10.1016/j.molcel.2020.08.012]

Structural Basis of Tail-Anchored Membrane Protein Biogenesis by the GET Insertase Complex

Fiorentino F.;
2020

Abstract

Membrane protein biogenesis faces the challenge of chaperoning hydrophobic transmembrane helices for faithful membrane insertion. The guided entry of tail-anchored proteins (GET) pathway targets and inserts tail-anchored (TA) proteins into the endoplasmic reticulum (ER) membrane with an insertase (yeast Get1/Get2 or mammalian WRB/CAML) that captures the TA from a cytoplasmic chaperone (Get3 or TRC40, respectively). Here, we present cryo-electron microscopy reconstructions, native mass spectrometry, and structure-based mutagenesis of human WRB/CAML/TRC40 and yeast Get1/Get2/Get3 complexes. Get3 binding to the membrane insertase supports heterotetramer formation, and phosphatidylinositol binding at the heterotetramer interface stabilizes the insertase for efficient TA insertion in vivo. We identify a Get2/CAML cytoplasmic helix that forms a "gating" interaction with Get3/TRC40 important for TA insertion. Structural homology with YidC and the ER membrane protein complex (EMC) implicates an evolutionarily conserved insertion mechanism for divergent substrates utilizing a hydrophilic groove. Thus, we provide a detailed structural and mechanistic framework to understand TA membrane insertion.
2020
cryo-EM; EMC; GET/TRC pathway; lipid binding; membrane proteins; native mass spectrometry; protein transport; tail anchor; YidC; Cell Line; Conserved Sequence; Evolution, Molecular; Humans; Membrane Proteins; Models, Molecular; Multiprotein Complexes; Phosphatidylinositols; Protein Binding; Protein Multimerization; Protein Stability; Protein Structure, Secondary; Saccharomyces cerevisiae; Saccharomyces cerevisiae Proteins
01 Pubblicazione su rivista::01a Articolo in rivista
Structural Basis of Tail-Anchored Membrane Protein Biogenesis by the GET Insertase Complex / Mcdowell, M. A.; Heimes, M.; Fiorentino, F.; Mehmood, S.; Farkas, A.; Coy-Vergara, J.; Wu, D.; Bolla, J. R.; Schmid, V.; Heinze, R.; Wild, K.; Flemming, D.; Pfeffer, S.; Schwappach, B.; Robinson, C. V.; Sinning, I.. - In: MOLECULAR CELL. - ISSN 1097-4164. - 80:1(2020), pp. 72-86. [10.1016/j.molcel.2020.08.012]
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Utilizza questo identificativo per citare o creare un link a questo documento: https://hdl.handle.net/11573/1561739
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