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Detection of Small GTPase Prenylation and GTP Binding Using Membrane Fractionation and GTPase-linked Immunosorbent Assay
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Biochemistry
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JoVE Journal Biochemistry
Detection of Small GTPase Prenylation and GTP Binding Using Membrane Fractionation and GTPase-linked Immunosorbent Assay

Detection of Small GTPase Prenylation and GTP Binding Using Membrane Fractionation and GTPase-linked Immunosorbent Assay

DOI:
10.3791/57646-v

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13:51 min

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November 11, 2018

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, , , , , , , ,
1Department of Human Anatomy and Cell Science, Rady Faculty of Health Sciences, Max Rady College of Medicine,University of Manitoba, 2Biology of Breathing Theme, Children Hospital Research Institute of Manitoba,University of Manitoba, 3Department of Biochemistry, School of Pharmacy and Pharmaceutical Sciences,Isfahan University of Medical Sciences, 4Division of Pulmonary, Critical Care, and Sleep Medicine, Department of Internal Medicine,Center for Comparative Respiratory Biology and Medicine, 5Department of Clinical Biochemistry, School of Medicine,Zahedan University of Medical Sciences, 6Department of Molecular Biology, School of Pharmacy with the Division of Laboratory Medicine in Sosnowiec,Medical University of Silesia, 7Centre de biophysique moléculaire – UPR 4301,Centre national de la recherche scientifique (CNRS) CS80054, 8LinkoCare Life Sciences AB, 9College of Nursing and Children’s Hospital Research Institute of Manitoba, Rady Faculty of Health Sciences,University of Manitoba, 10Health Policy Research Center, Institute of Health,Shiraz University of Medical Sciences

Chapters

  • 00:00Title
  • 01:39Determination of RhoA Localization Using Membrane/Cytosol Fractionation
  • 06:05Measurement of RhoA GTP Load using Small G-protein Activation Assay
  • 12:37Results
  • 13:09Conclusion

Summary

Automatic Translation

Automatic Translation

Here we describe a protocol to investigate the prenylation and guanosine-5'-triphosphate (GTP)-loading of Rho GTPase. This protocol consists of two detailed methods, namely membrane fractionation and a GTPase-linked immunosorbent assay. The protocol can be used for measuring the prenylation and GTP loading of different other small GTPases.

Tags

Small GTPaseRho GTPasePrenylationGTP BindingMembrane FractionationGTPase-linked Immunosorbent AssayGlioblastoma Cell LinesSimvastatinSubcellular FractionationRhoAPost-translational ModificationCell FateCellular ResponseCellular MotilityCell CycleProtein RegulationEDTAUltracentrifugationCytoplasmic FractionMembrane Fraction

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