Reconstitution and transport analysis of eukaryotic transporters in the post-genomic era

Hiroshi Omote, Yoshinori Moriyama

Research output: Chapter in Book/Report/Conference proceedingChapter

Abstract

Measuring transport activity through reconstituted proteoliposomes is a key technique to resolve numerous problems found in the traditional methods. The system includes overexpression, purification, and reconstitution of transporters. Mixing of purified transporter with lipid and dilution below the critical micelle concentration result in rapid generation of proteoliposomes. Incubation of proteoliposomes in the presence of a driving force initiates substrate uptake. After starting the reaction, samples are passed through a gel filtration column to separate proteoliposomes from the reaction mixture. Here, we describe step-by-step procedures for such reconstitution assays.

Original languageEnglish
Title of host publicationMethods in Molecular Biology
PublisherHumana Press Inc.
Pages343-352
Number of pages10
Volume1700
DOIs
Publication statusPublished - Jan 1 2018

Publication series

NameMethods in Molecular Biology
Volume1700
ISSN (Print)1064-3745

Keywords

  • Driving force
  • Kinetics
  • Proteoliposome
  • Purification
  • Reconstitution
  • Transporter

ASJC Scopus subject areas

  • Molecular Biology
  • Genetics

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  • Cite this

    Omote, H., & Moriyama, Y. (2018). Reconstitution and transport analysis of eukaryotic transporters in the post-genomic era. In Methods in Molecular Biology (Vol. 1700, pp. 343-352). (Methods in Molecular Biology; Vol. 1700). Humana Press Inc.. https://doi.org/10.1007/978-1-4939-7454-2_19