Advertisement

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
Article
  • 114 Downloads

Abstract

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.

Keywords

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

Abbreviations

PM

Plasma membrane

MPs

Membrane proteins

PMPs

Plasma membrane proteins

PMAPs

Plasma membrane-associated proteins

PBS

Phosphate-buffered saline

TBS

Tris-buffered saline

HB

Homogenization buffer

CHO

Chinese Hamster Ovary

MS

Mass spectrometry

IEF

Isoelectric focusing

2-DE

Two-dimensional electrophoresis

16-BAC/SDS-PAGE

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

LC–MS/MS

Liquid chromatography-mass spectrometry

Notes

Acknowledgements

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)

References

  1. Arnold T, Linke D (2007) Phase separation in the isolation and purification of membrane proteins. Biotechniques 43(427–30):432 (434 passim) Google Scholar
  2. Baharvand H, Fathi A, van Hoof D, Salekdeh GH (2007) Concise review: trends in stem cell proteomics. Stem Cells 25:1888–1903PubMedGoogle Scholar
  3. Blonder J, Conrads TP, Yu LR, Terunuma A, Janini GM, Issaq HJ, Vogel JC, Veenstra TD (2004) A detergent- and cyanogen bromide-free method for integral membrane proteomics: application to halobacterium purple membranes and the human epidermal membrane proteome. Proteomics 4:31–45PubMedGoogle Scholar
  4. Bordier C (1981) Phase separation of integral membrane proteins in Triton X-114 solution. J Biol Chem 256:1604–1607PubMedGoogle Scholar
  5. Bradford MM (1976) A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein-dye binding. Anal Biochem 72:248–254PubMedPubMedCentralGoogle Scholar
  6. Busch G, Hoder D, Reutter W, Tauber R (1989) Selective isolation of individual cell surface proteins from tissue culture cells by a cleavable biotin label. Eur J Cell Biol 50:257–262PubMedGoogle Scholar
  7. Chevalier F (2010) Highlights on the capacities of “gel-based” proteomics. Proteome Sci 8:23PubMedPubMedCentralGoogle Scholar
  8. Cordwell SJ, Thingholm TE (2010) Technologies for plasma membrane proteomics. Proteomics 10:611–627PubMedGoogle Scholar
  9. deBlaquiere J, Burgess AW (1999) Affinity purification of plasma membranes. J Biomol Tech 10:64–71PubMedPubMedCentralGoogle Scholar
  10. Donoghue PM, Hughes C, Vissers JP, Langridge JI, Dunn MJ (2008) Nonionic detergent phase extraction for the proteomic analysis of heart membrane proteins using label-free LC-MS. Proteomics 8:3895–3905PubMedGoogle Scholar
  11. Ellerbroek SM, Wu YI, Overall CM, Stack MS (2001) Functional interplay between type I collagen and cell surface matrix metalloproteinase activity. J Biol Chem 276:24833–24842PubMedGoogle Scholar
  12. Foster LJ, Zeemann PA, Li C, Mann M, Jensen ON, Kassem M (2005) Differential expression profiling of membrane proteins by quantitative proteomics in a human mesenchymal stem cell line undergoing osteoblast differentiation. Stem Cells 23:1367–1377PubMedGoogle Scholar
  13. Giansanti P, Tsiatsiani L, Low TY, Heck AJ (2016) Six alternative proteases for mass spectrometry-based proteomics beyond trypsin. Nat Protoc 11:993–1006PubMedGoogle Scholar
  14. Gilmore JM, Washburn MP (2010) Advances in shotgun proteomics and the analysis of membrane proteomes. J Proteom 73:2078–2091Google Scholar
  15. Gu B, Zhang J, Wu Y, Zhang X, Tan Z, Lin Y, Huang X, Chen L, Yao K, Zhang M (2011) Proteomic analyses reveal common promiscuous patterns of cell surface proteins on human embryonic stem cells and sperms. PLoS ONE 6:e19386PubMedPubMedCentralGoogle Scholar
  16. Hagner-McWhirter A, Winkvist M, Bourin S, Marouga R (2008) Selective labelling of cell-surface proteins using CyDye DIGE fluor minimal dyes. J Vis Exp 21:e945Google Scholar
  17. Hartinger J, Stenius K, Hogemann D, Jahn R (1996) 16-BAC/SDS-PAGE: a two-dimensional gel electrophoresis system suitable for the separation of integral membrane proteins. Anal Biochem 240:126–133PubMedGoogle Scholar
  18. Helbig AO, Heck AJ, Slijper M (2010) Exploring the membrane proteome–challenges and analytical strategies. J Proteom 73:868–878Google Scholar
  19. Hongsachart P, Sinchaikul S, Phutrakul S, Wongkham W, Chen S (2008) Comparative membrane extraction methods for identifying membrane proteome of SW900 squamous lung cancer cell line. Chiang Mai J Sci 35:467–482Google Scholar
  20. Islinger M, Weber G (2008) Free flow isoelectric focusing: a method for the separation of both hydrophilic and hydrophobic proteins of rat liver peroxisomes. Methods Mol Biol 432:199–215PubMedGoogle Scholar
  21. Jang JH, Hanash S (2003) Profiling of the cell surface proteome. Proteomics 3:1947–1954PubMedGoogle Scholar
  22. Josic D, Clifton JG (2007) Mammalian plasma membrane proteomics. Proteomics 7:3010–3029PubMedGoogle Scholar
  23. Kjeldsen L, Sengelov H, Borregaard N (1999) Subcellular fractionation of human neutrophils on Percoll density gradients. J Immunol Methods 232:131–143PubMedGoogle Scholar
  24. Laemmli UK (1970) Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature 227:680–685Google Scholar
  25. Lee SK, Kim Y, Kim SS, Lee JH, Cho K, Lee SS, Lee ZW, Kwon KH, Kim YH, Suh-Kim H, Yoo JS, Park YM (2009) Differential expression of cell surface proteins in human bone marrow mesenchymal stem cells cultured with or without basic fibroblast growth factor containing medium. Proteomics 9:4389–4405PubMedGoogle Scholar
  26. Li Q, Harraz MM, Zhou W, Zhang LN, Ding W, Zhang Y, Eggleston T, Yeaman C, Banfi B, Engelhardt JF (2006) Nox2 and Rac1 regulate H2O2-dependent recruitment of TRAF6 to endosomal interleukin-1 receptor complexes. Mol Cell Biol 26:140–154PubMedPubMedCentralGoogle Scholar
  27. Lilley KS, Friedman DB (2004) All about DIGE: quantification technology for differential-display 2D-gel proteomics. Expert Rev Proteom 1:401–409Google Scholar
  28. Luche S, Santoni V, Rabilloud T (2003) Evaluation of nonionic and zwitterionic detergents as membrane protein solubilizers in two-dimensional electrophoresis. Proteomics 3:249–253PubMedGoogle Scholar
  29. Lund R, Leth-Larsen R, Jensen ON, Ditzel HJ (2009) Efficient isolation and quantitative proteomic analysis of cancer cell plasma membrane proteins for identification of metastasis-associated cell surface markers. J Proteome Res 8:3078–3090PubMedGoogle Scholar
  30. Mathias RA, Chen YS, Kapp EA, Greening DW, Mathivanan S, Simpson RJ (2011) Triton X-114 phase separation in the isolation and purification of mouse liver microsomal membrane proteins. Methods 54:396–406PubMedGoogle Scholar
  31. Morre DJ, Morre DM (1989) Preparation of mammalian plasma membranes by aqueous two-phase partition. Biotechniques 7:946–948 (950–954, 956–958) PubMedGoogle Scholar
  32. Nunomura K, Nagano K, Itagaki C, Taoka M, Okamura N, Yamauchi Y, Sugano S, Takahashi N, Izumi T, Isobe T (2005) Cell surface labeling and mass spectrometry reveal diversity of cell surface markers and signaling molecules expressed in undifferentiated mouse embryonic stem cells. Mol Cell Proteom 4:1968–1976Google Scholar
  33. Orsburn BC, Stockwin LH, Newton DL (2011) Challenges in plasma membrane phosphoproteomics. Expert Rev Proteom 8:483–494Google Scholar
  34. Ozgul S, Kasap M, Akpinar G, Kanli A, Guzel N, Karaosmanoglu K, Baykal AT, Iseri P (2015) Linking a compound-heterozygous Parkin mutant (Q311R and A371T) to Parkinson’s disease by using proteomic and molecular approaches. Neurochem Int 85–86:1–13PubMedGoogle Scholar
  35. Ozlu N, Monigatti F, Renard BY, Field CM, Steen H, Mitchison TJ, Steen JJ (2010) Binding partner switching on microtubules and aurora-B in the mitosis to cytokinesis transition. Mol Cell Proteom 9:336–350Google Scholar
  36. Pionneau C, Canelle L, Bousquet J, Hardouin J, Bigeard J, Caron M (2005) Proteomic analysis of membrane-associated proteins from the breast cancer cell line MCF7. Cancer Genom Proteom 2:199–208Google Scholar
  37. Prive GG (2007) Detergents for the stabilization and crystallization of membrane proteins. Methods 41:388–397PubMedGoogle Scholar
  38. Qoronfleh MW, Benton B, Ignacio R, Kaboord B (2003) Selective enrichment of membrane proteins by partition phase separation for proteomic studies. J Biomed Biotechnol 2003:249–255PubMedPubMedCentralGoogle Scholar
  39. Rabilloud T (2009) Membrane proteins and proteomics: love is possible, but so difficult. Electrophoresis 30(Suppl 1):S174–S180PubMedGoogle Scholar
  40. Rabilloud T, Chevallet M, Luche S, Lelong C (2010) Two-dimensional gel electrophoresis in proteomics: past, present and future. J Proteom 73:2064–2077Google Scholar
  41. Rawlings AE (2016) Membrane proteins: always an insoluble problem? Biochem Soc Trans 44:790–795PubMedPubMedCentralGoogle Scholar
  42. Sabarth N, Lamer S, Zimny-Arndt U, Jungblut PR, Meyer TF, Bumann D (2002) Identification of surface proteins of Helicobacter pylori by selective biotinylation, affinity purification, and two-dimensional gel electrophoresis. J Biol Chem 277:27896–27902PubMedGoogle Scholar
  43. Santoni V, Rabilloud T, Doumas P, Rouquie D, Mansion M, Kieffer S, Garin J, Rossignol M (1999) Towards the recovery of hydrophobic proteins on two-dimensional electrophoresis gels. Electrophoresis 20:705–711PubMedGoogle Scholar
  44. Santoni V, Molloy M, Rabilloud T (2000) Membrane proteins and proteomics: un amour impossible? Electrophoresis 21:1054–1070PubMedGoogle Scholar
  45. Scheurer SB, Rybak JN, Roesli C, Brunisholz RA, Potthast F, Schlapbach R, Neri D, Elia G (2005) Identification and relative quantification of membrane proteins by surface biotinylation and two-dimensional peptide mapping. Proteomics 5:2718–2728PubMedGoogle Scholar
  46. Schiffer E, Mischak H, Novak J (2006) High resolution proteome/peptidome analysis of body fluids by capillary electrophoresis coupled with MS. Proteomics 6:5615–5627PubMedGoogle Scholar
  47. Shin BK, Wang H, Yim AM, Le Naour F, Brichory F, Jang JH, Zhao R, Puravs E, Tra J, Michael CW, Misek DE, Hanash SM (2003) Global profiling of the cell surface proteome of cancer cells uncovers an abundance of proteins with chaperone function. J Biol Chem 278:7607–7616PubMedGoogle Scholar
  48. Sidibe A, Yin X, Tarelli E, Xiao Q, Zampetaki A, Xu Q, Mayr M (2007) Integrated membrane protein analysis of mature and embryonic stem cell-derived smooth muscle cells using a novel combination of CyDye/biotin labeling. Mol Cell Proteom 6:1788–1797Google Scholar
  49. Simpson DC, Smith RD (2005) Combining capillary electrophoresis with mass spectrometry for applications in proteomics. Electrophoresis 26:1291–1305PubMedGoogle Scholar
  50. Smolders K, Lombaert N, Valkenborg D, Baggerman G, Arckens L (2015) An effective plasma membrane proteomics approach for small tissue samples. Sci Rep 5:10917PubMedPubMedCentralGoogle Scholar
  51. Tan S, Tan HT, Chung MC (2008) Membrane proteins and membrane proteomics. Proteomics 8:3924–3932PubMedGoogle Scholar
  52. Tanford C, Reynolds JA (1976) Characterization of membrane proteins in detergent solutions. Biochim Biophys Acta 457:133–170PubMedGoogle Scholar
  53. Tauber R, Reutter W (1978) Degradation of fucoproteins and sialoproteins in the plasma membrane of normal and regenerating liver. FEBS Lett 87:135–138PubMedGoogle Scholar
  54. Tsiatsiani L, Heck AJ (2015) Proteomics beyond trypsin. FEBS J 282:2612–2626PubMedGoogle Scholar
  55. Vit O, Petrak J (2016) Integral membrane proteins in proteomics. How to break open the black box? J Proteomics 153:8–20PubMedGoogle Scholar
  56. Wallin E, von Heijne G (1998) Genome-wide analysis of integral membrane proteins from eubacterial, archaean, and eukaryotic organisms. Protein Sci 7:1029–1038PubMedPubMedCentralGoogle Scholar
  57. Wenge B, Bonisch H, Grabitzki J, Lochnit G, Schmitz B, Ahrend MH (2008) Separation of membrane proteins by two-dimensional electrophoresis using cationic rehydrated strips. Electrophoresis 29:1511–1517PubMedGoogle Scholar
  58. Wu C, Lai CF, Mobley WC (2001) Nerve growth factor activates persistent Rap1 signaling in endosomes. J Neurosci 21:5406–5416PubMedPubMedCentralGoogle Scholar
  59. Wu CC, MacCoss MJ, Howell KE, Yates JR 3rd (2003) A method for the comprehensive proteomic analysis of membrane proteins. Nat Biotechnol 21:532–538PubMedGoogle Scholar
  60. Zhao Y, Zhang W, Kho Y (2004) Proteomic analysis of integral plasma membrane proteins. Anal Chem 76:1817–1823PubMedGoogle Scholar

Copyright information

© 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

Personalised recommendations