Advertisement

Comparative Haematology International

, Volume 1, Issue 3, pp 155–160 | Cite as

Electrophoretic analysis of erythrocyte membrane proteins and glycoproteins from different species

  • R. N. Barker
Original articles

Abstract

The erythrocyte membrane proteins and glycoproteins of man, rat, mouse, sheep and dog were analysed by sodium dodecyl sulphate polyacrylamide gel electrophoresis (SDS-PAGE), using a discontinuous buffer system. Considerable similarities between the species were observed in the pattern of protein bands seen when gels were stained with Coomassie Blue. Equivalents to human Bands 1, 2, 3, 4.2, 5 and 8 appeared to be present in the rat, mouse, sheep and dog, and Band 4.1 was identified as a closely spaced doublet in all species except the rat.

When RBC membranes were stained with periodic acid-Schiff (PAS) after SDS-PAGE analysis, glycoproteins equivalent to human glycophorins were identified in all the species studied. However, in contrast to the overall similarity of the protein patterns, the number, relative staining intensity and apparent molecular masses of the PAS-stained bands differed between species.

The silver stain was assessed in the detection of RBC membrane proteins in polyacrylamide gels, and found to be more sensitive than Coomassie Blue. The technique also stained many of the PAS-positive glycophorins as diffuse orange zones, which could be distinguished from the darker protein bands by their differential colouration.

In view of the interspecies variation in the glycophorins after SDS-PAGE, it is suggested that, unlike the membrane proteins, their functions do not require a conserved structure.

Keywords

Erythrocyte Membrane proteins Membrane glycoproteins Comparative study Gel electrophoresis 

Preview

Unable to display preview. Download preview PDF.

Unable to display preview. Download preview PDF.

References

  1. Anstee DJ, Mawby WJ, Tanner MJA (1979) Abnormal blood-groupSs-active sialoglycoproteins in the membrane of Miltenberger class III, IV and V human erythrocytes. Biochem J 183:193–203Google Scholar
  2. Barker RN, Gruffydd-Jones TJ, Stokes CR, Elson CJ (1991) Identification of autoantigens in canine autoimmune haemolytic anaemia. Clin Exp Immunol [in press]Google Scholar
  3. Bennett V (1985) The membrane skeleton of human erythrocytes and its implications for more complex cells. Ann Rev Biochem 54:273–304Google Scholar
  4. 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 72:248–254Google Scholar
  5. Brown WRA, Barclay AN, Sunderland CA, Williams AF (1981) Identification of glycophorin-like molecule at the cell surface of rat thymocytes. Nature 289:456–460Google Scholar
  6. Burk RR, Eschenbruch M, Leuthard P, Steck G (1983) Sensitive detection of proteins and peptides in polyacrylamide gels after formaldehyde fixation. Methods Enzymol 91:247–254Google Scholar
  7. Cabantchik ZI, Knauf PA, Rothstein A (1978) The anion transport system of the red blood cell. The role of membrane protein evaluated by the use of probes. Biochim Biophys Acta 515:239–302Google Scholar
  8. Carraway KL, Kobylka D (1970) Comparative studies of erythrocyte membranes by gel electrophoresis. Biochim Biophys Acta 219:238–241Google Scholar
  9. Dodge JT, Mitchell C, Hanahan DJ (1963) The preparation and chemical characteristics of hemoglobin-free ghosts of human erythrocytes. Arch Biochem Biophys 100:119–130Google Scholar
  10. Dzandu JK, Deh ME, Barratt DL, Wise GE (1984) Detection of erythrocyte membrane proteins, sialoglycoproteins and lipids in the same polyacrylamide gel using a double-staining technique. Proc Natl Acad Sci 81:1733–1737Google Scholar
  11. Edge ASB, Webber P (1981) Purification and characterization of the major sialoglycoproteins of the rat erythrocyte membrane. Arch Biochem Biophys 209:697–705Google Scholar
  12. Fairbanks G, Steck TL, Wallach DFH (1971) Electrophoretic analysis of the major polypeptides of the human erythrocyte membrane. Biochemistry 10:2606–2617Google Scholar
  13. Fukada M, Fukada MN (1984) Cell surface glycoproteins and carbo hydrate antigens in development and differentiation of human erythroid cells. In: Ivatt RJ (ed.) The biology of glycoproteins. Plenum, New York, pp 183–234Google Scholar
  14. Gahmberg CG, Myllyla G, Leikola J, Pirkola A, Nordling S (1976) Absence of the major sialoglycoprotein in the membrane of human En(a-) erythrocytes and increased glycosylation of band 3. J Biol Chem 251:6108–6116Google Scholar
  15. Glossman H, Neville DM (1971) Glycoproteins of cell surfaces. A comparative study of 3 different cell surfaces of the rat. J Biol Chem 246:6339–6346Google Scholar
  16. Hamaguchi H, Cleve, H (1972) Solubilization and comparative analysis of mammalian erythrocyte membrane glycoproteins. Biochem Biophys Res Commun 47:459–464Google Scholar
  17. Hitsumoto Y, Hinemo T, Utsumi S (1984) Induction of tolerance by haptenated liposomes carrying mouse erythrocyte membrane glycoprotein. Immunology 53:847–854Google Scholar
  18. Kobylka D, Khettry A, Shin BC, Carraway KL (1972) Proteins and glycoproteins of the erythrocyte membrane. Arch Biochem Biophys 148:475–487Google Scholar
  19. Laemmli UK (1970) Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature 227:680–685Google Scholar
  20. Laing P, Appleby LA, Culbert EJ, Elson CJ (1988) Antigenicity of rat erythrocyte glycophorins. Immunol Lett 18:125–128Google Scholar
  21. Laing P, Parkar BA, Culbert EJ, Watt GJ, Elson CJ (1987) Identification of rat erythrocyte antigens with a new non-radioactive immunoprecipitation technique. Scand J Immunol 25:613–620Google Scholar
  22. Lenard J (1970) Protein components of erythrocyte membranes from different animal species. Biochemistry 9:5037–5040Google Scholar
  23. Maddy AH (1982) The solubilization of erythrocyte membrane proteins. In: Ellory JC, Young JD (eds.) Red cell membranes — a methodological approach. Academic Press, London, pp 43–66Google Scholar
  24. Marchesi VT, Furthmayr H, Tomita M (1976) The red cell membrane. Ann Rev Biochem 45:667–698Google Scholar
  25. Mueller TJ, Dow AW, Morrison M (1976) Heterogeneity of the sialoglycoproteins of the normal human erythrocyte membrane. Biochem Biophys Res Commun 72:94–99Google Scholar
  26. Mueller TJ, Morrison M (1980) Glycoconnectin (PAS 2), a component of the cytoskeleton of the human erythrocyte membrane. J Cell Biol 87:202A (abstr.)Google Scholar
  27. Noeman SA, Gill TJ III, Misra DN (1980) Characterization of the T antigen and T agglutinin in inbred rats. Immunology 39:219–229Google Scholar
  28. Ralston GB (1975) Proteins of the camel erythrocyte membrane. Biochim Biophys Acta 401:83–94Google Scholar
  29. Sarris AH, Palade GE (1979) The sialoglycoproteins of murine erythrocyte ghosts. J Biol Chem 254:6724–6731Google Scholar
  30. Sarris AH, Palade GE (1982) Isolation and partial characterization of the sialoglycoproteins of murine erythroid cells. J Cell Biol 93:583–590Google Scholar
  31. Segrest JP, Jackson RL, Andrews EP, Marchesi VT (1971) Human erythrocyte membrane glycoprotein: a re-evaluation of the molecular weight as determined by SDS polyacrylamide gel electrophoresis. Biochem Biophys Res Commun 44:390–395Google Scholar
  32. Steck TL (1974) The organization of proteins in the human red blood cell membrane. J Cell Biol 62:1–19Google Scholar
  33. Thompson S, Maddy AH (1982) Gel electrophoresis of erythrocyte membrane proteins. In: Ellory JC, Young JD (eds.) Red cell membranes — a methodological approach. Academic Press, London, pp 67–93Google Scholar

Copyright information

© Springer-Verlag London Limited 1991

Authors and Affiliations

  • R. N. Barker
    • 1
  1. 1.Department of Pathology and MicrobiologyUniversity of Bristol, Medical School, University WalkBristolUK

Personalised recommendations