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Related Concept Videos

GPI Anchoring of Proteins in the ER Membrane01:29

GPI Anchoring of Proteins in the ER Membrane

GPI-anchoring is a post-translational, reversible protein modification that is ubiquitous in eukaryotes. Such proteins are primarily present on the exoplasmic leaflet of the plasma membrane.
GPI-anchor structure
A sequence of 11 enzymatic reactions results in the synthesis of the complete GPI anchor consisting of a hydrophobic and a hydrophilic portion. The hydrophobic portion comprises phosphatidylinositol, while the hydrophilic part comprises polar groups like phosphoethanolamine,...
Lipids as Anchors01:32

Lipids as Anchors

In the plasma membrane, the lipids forming the bilayer can also act as an anchor to tether proteins to the membrane. The three main types of lipid anchors found in eukaryotes are – prenyl groups, fatty acyl groups, and glycosylphosphatidylinositol or GPI groups. Prenyl and fatty acyl groups act as anchors on the cytosolic surface of the membrane, whereas GPI anchors proteins on the extracellular side.
The carboxy-terminal of most of the prenylated proteins, such as Ras proteins, contains the...
Phosphoinositides and PIPs01:42

Phosphoinositides and PIPs

Phosphoinositides are a group of phospholipids containing a glycerol backbone with two fatty acid chains and a phosphate attached to a myoinositol sugar ring. The inositol head group extends into the cytoplasm, where it is modified by adding phosphate groups to form phosphatidylinositol phosphates or PIPs.
Different phosphoinositides are synthesized and recruited on the cytosolic face of the plasma membrane. The localization of specific phosphoinositides concentrated in separate membrane...

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Related Experiment Video

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Transient Expression and Cellular Localization of Recombinant Proteins in Cultured Insect Cells
12:09

Transient Expression and Cellular Localization of Recombinant Proteins in Cultured Insect Cells

Published on: April 20, 2017

An efficient method to express GPI-anchor proteins in insect cells.

Hosam Shams-Eldin1, Nahid Azzouz, Sebastian Niehus

  • 1Medizinisches Zentrum für Hygiene und Medizinische Mikrobiologie, Philipps-Universität Marburg, Hans-Meerwein-Str. 2, 35043 Marburg, Germany.

Biochemical and Biophysical Research Communications
|November 22, 2007
PubMed
Summary

This study introduces a novel method for expressing glycosylphosphatidylinositol (GPI) anchor proteins in insect cells. By overexpressing a key enzyme, researchers overcome baculovirus infection

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Optimizing the Genetic Incorporation of Chemical Probes into GPCRs for Photo-crosslinking Mapping and Bioorthogonal Chemistry in Live Mammalian Cells
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Optimizing the Genetic Incorporation of Chemical Probes into GPCRs for Photo-crosslinking Mapping and Bioorthogonal Chemistry in Live Mammalian Cells
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Optimizing the Genetic Incorporation of Chemical Probes into GPCRs for Photo-crosslinking Mapping and Bioorthogonal Chemistry in Live Mammalian Cells

Published on: April 9, 2018

Area of Science:

  • Biochemistry
  • Cell Biology
  • Molecular Biology

Background:

  • Glycosylphosphatidylinositols (GPIs) are essential glycolipids anchoring proteins to eukaryotic cell surfaces.
  • Expressing parasitic protozoa surface antigens in heterologous systems requires consideration of GPI anchor synthesis.
  • Baculovirus infection significantly inhibits endogenous GPI-anchor synthesis in insect cells.

Purpose of the Study:

  • To develop a method for efficient expression of GPI-anchor proteins in insect cells.
  • To overcome the limitations imposed by baculovirus-induced inhibition of GPI biosynthesis.

Main Methods:

  • Utilized a supplementary baculovirus construct designed for overexpression.
  • The construct overexpressed N-acetylglucosaminyl phosphatidylinositol de-N-acetylase, a crucial enzyme in the GPI pathway.
  • Insect cells were infected with this construct to enhance GPI anchor protein expression.

Main Results:

  • Successfully demonstrated a method to express GPI-anchor proteins in insect cells.
  • The supplementary baculovirus construct effectively counteracted the inhibitory effects of infection on GPI synthesis.
  • Achieved significant restoration of GPI anchor protein expression levels.

Conclusions:

  • The developed method enables robust expression of GPI-anchor proteins in insect cells.
  • This approach is vital for studies involving surface antigens from parasitic protozoa.
  • Overexpression of key GPI biosynthetic enzymes can restore protein expression despite viral interference.