C-tail-anchored (TA) proteins often require a transmembrane domain of moderate hydrophobicity to maintain their endoplasmic reticulum residence, but the suggested role of protein–lipid interactions in this phenomenon has not been established. Here, we studied the interaction of TA proteins with lipids by differential scanning calorimetry by using a model system consisting of liposomes embedding either of two forms of cytochrome b5: the endoplasmic reticulum-resident wild-type (b5wt) and a mutant thereof (b5ext), that contains five extra nonpolar amino acids in its transmembrane domain and, therefore, reaches the plasma membrane. The proteins were incorporated into liposomes of palmitoyloleyl- phosphatidylcholine (POPC) or POPC mixed with either distearoyl-phosphatidylserine (DSPS), palmitoyl-oleyl-phosphatidylserine (POPS), distearoyl-phosphatidylcholine (DSPC), or C16- ceramide (CER). POPC liposomes displayed a single thermotropic transition centered at 3.4°C. When present, the second lipid formed a domain within the POPC bilayer, as indicated by the appearance of an additional peak. This peak was centered at temperatures close to 0°C in the case of liposomes containing 10% CER, DSPS, and POPS and at 23°C in the case of DSPC, likely reflecting a higher degree of molecular packing for DSPC domains. In DSPS POPC, POPS POPC, or CER POPC, but not in DSPC POPC liposomes, the insertion of b5wt increased, whereas b5ext decreased, the relative contribution to the total enthalpy of the higher temperature, phase-separated component. These results were confirmed with fluorescence measurements by using pyrenelabeled phospholipids. The dissimilar interaction with lipids of these two differently localized TA proteins could have implications for their intracellular sorting.

Ceppi, P., Colombo, S., Francolini, M., Raimondo, F., Borgese, N., & Masserini, M. (2005). Two tail-anchored protein variants, differing in transmembrane domain length and intracellular sorting, interact differently with lipids. PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA, 102, 16269-16274 [10.1073/pnas.0508157102].

Two tail-anchored protein variants, differing in transmembrane domain length and intracellular sorting, interact differently with lipids

RAIMONDO, FRANCESCA;MASSERINI, MASSIMO ERNESTO
2005

Abstract

C-tail-anchored (TA) proteins often require a transmembrane domain of moderate hydrophobicity to maintain their endoplasmic reticulum residence, but the suggested role of protein–lipid interactions in this phenomenon has not been established. Here, we studied the interaction of TA proteins with lipids by differential scanning calorimetry by using a model system consisting of liposomes embedding either of two forms of cytochrome b5: the endoplasmic reticulum-resident wild-type (b5wt) and a mutant thereof (b5ext), that contains five extra nonpolar amino acids in its transmembrane domain and, therefore, reaches the plasma membrane. The proteins were incorporated into liposomes of palmitoyloleyl- phosphatidylcholine (POPC) or POPC mixed with either distearoyl-phosphatidylserine (DSPS), palmitoyl-oleyl-phosphatidylserine (POPS), distearoyl-phosphatidylcholine (DSPC), or C16- ceramide (CER). POPC liposomes displayed a single thermotropic transition centered at 3.4°C. When present, the second lipid formed a domain within the POPC bilayer, as indicated by the appearance of an additional peak. This peak was centered at temperatures close to 0°C in the case of liposomes containing 10% CER, DSPS, and POPS and at 23°C in the case of DSPC, likely reflecting a higher degree of molecular packing for DSPC domains. In DSPS POPC, POPS POPC, or CER POPC, but not in DSPC POPC liposomes, the insertion of b5wt increased, whereas b5ext decreased, the relative contribution to the total enthalpy of the higher temperature, phase-separated component. These results were confirmed with fluorescence measurements by using pyrenelabeled phospholipids. The dissimilar interaction with lipids of these two differently localized TA proteins could have implications for their intracellular sorting.
Articolo in rivista - Articolo scientifico
Scientifica
cytochrome b5, differential scanning calorimetry, endoplasmic reticulum, lipid domains, liposomes
English
16269
16274
Ceppi, P., Colombo, S., Francolini, M., Raimondo, F., Borgese, N., & Masserini, M. (2005). Two tail-anchored protein variants, differing in transmembrane domain length and intracellular sorting, interact differently with lipids. PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA, 102, 16269-16274 [10.1073/pnas.0508157102].
Ceppi, P; Colombo, S; Francolini, M; Raimondo, F; Borgese, N; Masserini, M
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Utilizza questo identificativo per citare o creare un link a questo documento: http://hdl.handle.net/10281/15089
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