The Journal of Membrane Biology

, Volume 252, Issue 6, pp 587–608 | Cite as

Comparative Proteomics Analysis of Four Commonly Used Methods for Identification of Novel Plasma Membrane Proteins

  • Kubra Karaosmanoglu Yoneten
  • Murat KasapEmail author
  • Gurler Akpinar
  • Aylin Kanli
  • Erdal Karaoz


Plasma membrane proteins perform a variety of important tasks in the cells. These tasks can be diverse as carrying nutrients across the plasma membrane, receiving chemical signals from outside the cell, translating them into intracellular action, and anchoring the cell in a particular location. When these crucial roles of plasma membrane proteins are considered, the need for their characterization becomes inevitable. Certain characteristics of plasma membrane proteins such as hydrophobicity, low solubility, and low abundance limit their detection by proteomic analyses. Here, we presented a comparative proteomics study in which the most commonly used plasma membrane protein enrichment methods were evaluated. The methods that were utilized include biotinylation, selective CyDye labeling, temperature-dependent phase partition, and density-gradient ultracentrifugation. Western blot analysis was performed to assess the level of plasma membrane protein enrichment using plasma membrane and cytoplasmic protein markers. Quantitative evaluation of the level of enrichment was performed by two-dimensional electrophoresis (2-DE) and benzyldimethyl-n-hexadecylammonium chloride/sodium dodecyl sulfate polyacrylamide gel electrophoresis (16-BAC/SDS-PAGE) from which the protein spots were cut and identified. Results from this study demonstrated that density-gradient ultracentrifugation method was superior when coupled with 16-BAC/SDS-PAGE. This work presents a valuable contribution and provides a future direction to the membrane sub-proteome research by evaluating commonly used methods for plasma membrane protein enrichment.


Plasma membrane proteins Membrane protein enrichment 2-DE 16-BAC/SDS-PAGE 



Plasma membrane


Membrane proteins


Plasma membrane proteins


Plasma membrane-associated proteins


Phosphate-buffered saline


Tris-buffered saline


Homogenization buffer


Chinese Hamster Ovary


Mass spectrometry


Isoelectric focusing


Two-dimensional electrophoresis


Benzyldimethyl-n-hexadecylammonium chloride/sodium dodecyl sulfate polyacrylamide gel electrophoresis


Liquid chromatography-mass spectrometry



This work was supported by The Scientific and Technological Research Council of Turkey (TUBITAK) under the Grant Number of 113S868.

Supplementary material

232_2019_84_MOESM1_ESM.tif (1010 kb)
Supplementary Figure: Flowchart depicting the steps of the study. Supplementary material 1 (TIFF 1009 kb)


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© Springer Science+Business Media, LLC, part of Springer Nature 2019

Authors and Affiliations

  1. 1.Biomedical Engineering Department, Technology FacultyKocaeli UniversityKocaeliTurkey
  2. 2.Department of Medical Biology, School of MedicineKocaeli UniversityKocaeliTurkey
  3. 3.Department of Histology and Embryology, School of MedicineIstinye UniversityIstanbulTurkey

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