The use of liposomes to study COPII- and COPI-coated vesicle formation and membrane protein sorting

Ken Matsuoka, Randy Schekman

Research output: Contribution to journalArticlepeer-review

27 Citations (Scopus)

Abstract

We have established systems that reconstitute the biogenesis of coated transport vesicles with liposomes made of pure lipids and purified coat proteins. Optimization of the lipid composition in the liposomes allowed the efficient binding of both coat protein I and coat protein II (COPII) coat subunits. Coated vesicles of approximately the size generated from biomembranes were detected and characterized by centrifugation analysis and electron microscopy. A variation of this budding reaction allowed us to measure the sorting of v-SNARE proteins into synthetic COPII vesicles. We developed a novel system to tether glutathione S-transferase (GST)-hybrid proteins to the surface of liposomes formulated with a glutathione-derivatized phospholipid. This system allowed us to detect the positive role of cytoplasmic domains of two v-SNARE proteins that are packaged into COPII vesicles. Therefore, both generation of coated vesicles and protein sorting into the vesicles can be reproduced with liposomes and purified proteins.

Original languageEnglish
Pages (from-to)417-428
Number of pages12
JournalMethods
Volume20
Issue number4
DOIs
Publication statusPublished - 2000
Externally publishedYes

All Science Journal Classification (ASJC) codes

  • Molecular Biology
  • Biochemistry, Genetics and Molecular Biology(all)

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