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Related Concept Videos

ATP Driven Pumps II: P-type Pumps01:34

ATP Driven Pumps II: P-type Pumps

6.8K
The P-type pumps are a large family of integral membrane transporter ATPases. They are divided into five major types based on substrate specificity, from I to V.
A typical P-type pump has three cytosolic domains: nucleotide-binding (N), phosphorylation (P), and activator (A) domains. These domains are connected to the membrane-spanning helices by short amino acid segments. ATP hydrolysis and covalent phosphoenzyme intermediate formation are crucial parts of the catalytic cycle. At the highly...
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ATP Driven Pumps I: An Overview01:27

ATP Driven Pumps I: An Overview

10.4K
ATP-driven pumps, also known as transport ATPases, are integral membrane proteins. They have binding sites for ATP located on the membrane's cytosolic side and the ion-conducting domain in the transmembrane region. These pumps use the free energy released from ATP hydrolysis to move the solutes across cell membranes against an electrochemical gradient.
There are four main types of ATP-driven pumps - P-type, V-type, F-type, and ABC transporter. All these pumps are of varying complexities and...
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ATP Driven Pumps III: V-type Pumps01:30

ATP Driven Pumps III: V-type Pumps

5.2K
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...
5.2K

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Related Experiment Video

Updated: Mar 28, 2026

Single Liposome Measurements for the Study of Proton-Pumping Membrane Enzymes Using Electrochemistry and Fluorescent Microscopy
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Single Liposome Measurements for the Study of Proton-Pumping Membrane Enzymes Using Electrochemistry and Fluorescent Microscopy

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Assaying P-Type ATPases Reconstituted in Liposomes.

Hans-Jürgen Apell1, Bojana Damnjanovic2

  • 1Department of Biology, University of Konstanz, 635, Universitätsstr. 10, Konstanz, 78464, Germany. h-j.apell@uni-konstanz.de.

Methods in Molecular Biology (Clifton, N.J.)
|December 24, 2015
PubMed
Summary
This summary is machine-generated.

Researchers reconstituted P-type ATPases, vital ion transporters, into liposomes. This method allows detailed study of their function, binding affinities, and transport properties using fluorescence dyes.

Keywords:
BioBeadsDetergentsDialysisElectrogenic ion transportLipidsMembrane potentialPump currentReconstitutionSubstrate dependenceVesiclesVoltage-sensitive fluorescence dyes

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Measuring In Vitro ATPase Activity for Enzymatic Characterization
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Area of Science:

  • Biochemistry
  • Molecular Biology
  • Biophysics

Background:

  • P-type ATPases are essential active ion transporters crucial for cellular function.
  • Reconstitution into unilamellar liposomes facilitates the study of their functional properties.
  • Understanding ion pump mechanisms requires analyzing substrate binding and transport kinetics.

Purpose of the Study:

  • To describe methods for reconstituting P-type ATPases into liposomes.
  • To detail the use of potential-sensitive fluorescence dyes for studying ion transport.
  • To analyze fluorescence signals for insights into P-type ATPase properties.

Main Methods:

  • Liposome reconstitution using dialysis.
  • Incorporation of Na,K-ATPase (rabbit kidney) and KdpFABC complex (E. coli).
  • Utilizing potential-sensitive fluorescence dyes to monitor ion transport.

Main Results:

  • Successful reconstitution of two distinct P-type ATPases.
  • Demonstration of fluorescence-based techniques for analyzing transport.
  • Quantification of substrate-binding affinities and ion pump current dependence on lipids and temperature.

Conclusions:

  • Liposome reconstitution is a valuable technique for P-type ATPase functional studies.
  • Fluorescence dye analysis provides detailed insights into ion transport mechanisms.
  • This approach enables comprehensive characterization of ion pump activity.