Springer Protocols: Abstract: Expression, Localization, and Topology of Fluorescently Tagged Plasma Membrane-Targeted Transmembrane Proteins

Expression, Localization, and Topology of Fluorescently Tagged Plasma Membrane-Targeted Transmembrane Proteins

By: Jeanne Shepshelovich², Koret Hirschberg²

Abstract

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Transmembrane proteins constitute a significant proportion of the total number of proteins encoded in eukaryotic genomes. These proteins are involved in countless processes required for cellular function and homeostasis. Mutations mapping within their genes form the cellular mechanisms for a variety of pathological conditions. The surface expression of polytopic proteins can be limited by their relatively slow folding in the endoplasmic reticulum (ER). This is especially evident in heterologous overexpression systems cite ch26:bib01 . On the other hand, expression of these proteins in endogenously expressing primary cell lines is generally a challenging task in itself. Here we present a comprehensive scheme to establish arrival at the plasma membrane (PM) of transiently expressed, fluorescently tagged transmembrane proteins and provide two experimental tools to determine whether a distinct domain is cytosolic or extracellular using fluorescence microscopy.

Affiliation(s): (2) Department of Pathology, Sackler School of Medicine, Tel-Aviv University, Tel Aviv, Israel

Book Title: Protein Targeting Protocols

Series: Methods in Molecular Biology | Volume: 390 | Pub. Date: May-21-2007 | Page Range: 393-403 | DOI: 10.1007/978-1-59745-466-7_26

Subject: Protein Science

Key Words: Green fluorescent protein - surface expression - transmembrane proteins - encoplasmic reticulum - Golgi apparatus - plasma membrane - saponin

References

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