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Related Concept Videos

ATP Driven Pumps III: V-type Pumps01:30

ATP Driven Pumps III: V-type Pumps

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V-type pumps are ATP-driven pumps found in the vacuolar membranes of plants, yeast, endosomal and lysosomal membranes of animal cells, plasma membranes of a few specialized eukaryotic cells, and some prokaryotes. They are also known as the V1Vo-ATPase, that couple ATP hydrolysis to transport protons against a concentration gradient.
The peripheral or cytosolic V1 domain with eight subunits is involved in ATP hydrolysis. The integral or transmembrane V0 domain containing at least five subunits...
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Protein Transport to the Outer Chloroplast Membrane01:11

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Chloroplast outer membrane proteins encoded by the nucleus are synthesized in the cytosol. Soon after synthesis, they bind cytosolic factors such as 14-3-3 protein and the Hsp70 chaperones that keep these precursors in an unfolded state until their translocation.
Two models describe the mechanism of precursor recognition and entry across the outer membrane through the TOC complex. Model 1 suggests the newly synthesized precursor binds to the TOC receptor 159 and forms a complex.
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Protein Transport to the Inner Chloroplast Membrane01:18

Protein Transport to the Inner Chloroplast Membrane

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Proteins targeted to the inner chloroplast membrane, or plastid proteins, are transported by two general pathways: the stop-transfer and the re-insertion or post-import pathways. Most plastid proteins carry N-terminal transit sequences and internal import sequences targeting it to the specific chloroplast subcompartment. Proteins targeted by the stop-transfer pathway have internal hydrophobic sequences that inhibit their translocation into the stroma. As a result, these precursors are arrested...
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Protein Transport to the Stroma01:24

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Chloroplasts are triple membrane structures with an outer membrane, an inner membrane, and a thylakoid membrane, each containing distinct metabolite transporters, membrane translocons, and enzymes. Appropriate sorting and translocating these proteins to their correct membrane systems is essential for chloroplast function.
Protein complexes called the translocon of the outer chloroplast membrane or TOC complex, and the translocon of the inner chloroplast membrane or TIC complex mediate the...
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The Apoplast and Symplast01:46

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Plant growth depends on its ability to take up water and dissolved minerals from the soil. The root system of every plant is equipped with the necessary tissues to facilitate the entry of water and solutes. The plant tissues involved in the transport of water and minerals have two major compartments - the apoplast and the symplast. The apoplast includes everything outside the plasma membrane of living cells and consists of cell walls, extracellular spaces, xylem, phloem, and tracheids. The...
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Introduction to Membrane Proteins01:16

Introduction to Membrane Proteins

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The cell membrane, or plasma membrane, is an ever-changing landscape. It is described as a fluid mosaic where various macromolecules are embedded in the phospholipid bilayer. Among the macromolecules are proteins. The protein content varies across cell types. For example, mitochondrial inner membranes contain ~76% protein content, while myelin contains ~18% protein content. Individual cells contain many types of membrane proteins—red blood cells contain over 50—and different cell...
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Updated: Feb 19, 2026

Metabolic Labeling and Membrane Fractionation for Comparative Proteomic Analysis of Arabidopsis thaliana Suspension Cell Cultures
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Analyzing the Vacuolar Membrane (Tonoplast) Proteome.

Miwa Ohnishi1, Katsuhisa Yoshida1, Tetsuro Mimura2

  • 1Department of Biology, Graduate School of Science, Kobe University, Rokkodai 1-1, Nada, Kobe, 657-8501, Japan.

Methods in Molecular Biology (Clifton, N.J.)
|November 1, 2017
PubMed
Summary

Researchers developed a simple method to isolate intact vacuoles and analyze vacuolar membrane (VM) proteins using proteomics. This advance aids in identifying novel tonoplast proteins and understanding vacuole function.

Keywords:
ArabidopsisDRM (detergent-resistant membrane)DSM (detergent-soluble membrane)MicrodomainPlant cellProteomeProteomicsTonoplastVacuolar membraneVacuole

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Area of Science:

  • Plant cell biology
  • Proteomics
  • Molecular biology

Background:

  • The vacuole is crucial for plant cells, with its membrane (tonoplast) housing vital transporters and receptors.
  • Identifying tonoplast proteins is essential for understanding vacuole function, but few have been characterized at the molecular level.
  • Current methods for studying tonoplast proteins are limited, hindering physiological role characterization.

Purpose of the Study:

  • To present a straightforward protocol for isolating intact vacuoles from Arabidopsis suspension cultures.
  • To enable comprehensive proteomic analysis of the vacuolar membrane (VM) fraction.
  • To facilitate the identification and characterization of a broader range of tonoplast proteins.

Main Methods:

  • Isolation of intact vacuoles from Arabidopsis suspension culture cells.
  • Subsequent proteomic analysis of the isolated vacuolar membrane (VM) fraction.
  • Utilizing mass spectrometry for accurate protein composition analysis of the VM.

Main Results:

  • A simplified method for obtaining pure vacuole isolates was established.
  • Proteomic analysis of the vacuolar membrane fraction was successfully performed.
  • The method provides a foundation for identifying numerous previously uncharacterized tonoplast proteins.

Conclusions:

  • The described method offers a reliable approach for vacuolar membrane proteome analysis.
  • This technique will significantly advance the identification of tonoplast proteins and their functions.
  • Enhanced understanding of vacuole biology is achievable through improved proteomic strategies.