Advertisement

Identification and Analysis of a Ca2+-Dependent Lactoferrin Receptor in Rat Liver

Lactoferrin Binds to the Asialoglycoprotein Receptor in a Galactose-Independent Manner
  • Douglas D. McAbee
  • David J. Bennatt
  • Yuan Yuan Ling
Part of the Advances in Experimental Medicine and Biology book series (AEMB, volume 443)

Summary

We identified a 45 kDa Ca2+-dependent Lf binding protein on rat hepatocytes. Dithiobis(sulfosuccimidylproprionate) (DTSSP)-crosslinked 125I-Lf to a 45 kDa adduct in a Ca2+-dependent manner on intact cells. The 125I-labeledcrosslinked complexes were absent when either surface-bound 125I-Lf was stripped prior to crosslinking or an excess of unlabeled Lf was included in the DTSSP reaction. Triton X-100 extracts of hepatocyte membrane ghosts were chromatographed on Lf-agarose, and a 45 kDa polypeptide (p45) was eluted by EGTA. Anti-p45 sera blocked vigorously 125I-Lf endocytosis to intact rat hepatocytes, confirming that p45 functions as the Ca2+-dependent Lf receptor on hepatocytes. Two tryptic fragments of p45 showed 100% identity with internal sequences (Leu121 →Lys 126 and Phe198-Lys220) of the major subunit (RHL- l) of the rat asialoglycoprotein receptor. Antisera against p45 and RHL-1 crossreacted equally well with each protein, and asialoorosomucoid blocked the binding of 125I-Lf to hepatocytes. We did not detect the minor subunits (RHL2/3) of the rat asialoglycoprotein receptor in p45 preparations from Triton X-100-extracts of hepatocytes, and 125I-Lf bound to immobilized RHL-1 but not to RHL-2/3. Exoglycosidases were used to remove terminally-exposed NeuNAc and α-and β-Gal from bovine Lf glycans, and lectin blotting confirmed that glycosidase-treated Lfs lacked detectable terminal Gal. Unexpectedly, deglycosylated Lf exhibited no loss in its ability to compete with unmodified Lf for binding to isolated hepatocytes. Moreover, β-lactose but not sucrose competed vigorously for 125I-Lf endocytosis by hepatocytes, indicating that Lf binds at or near the carbohydrate-recognition domain of RHL-l. We conclude that RHL-1 is the Ca2+-dependent Lf receptor on hepatocytes and that it binds Lf in a Gal-independent manner.

Keywords

Asialoglycoprotein Receptor Major Subunit Lectin Blotting Unbind Ligand Share Amino Acid Sequence 
These keywords were added by machine and not by the authors. This process is experimental and the keywords may be updated as the learning algorithm improves.

Preview

Unable to display preview. Download preview PDF.

Unable to display preview. Download preview PDF.

References

  1. 1.
    Huettinger, M, Retzek H, Hermann M, Goldenberg H. (1992) J. Biol. Chem. 267: 18551–18557.Google Scholar
  2. 2.
    Mellinger, M, Hamner M, Szakmary KA, Steinbock F, Scheiber B, Goldenberg H, Huettinger M. (1995) FEBS Lett. 360: 70–74.CrossRefGoogle Scholar
  3. 3.
    Hu, WL, Regoeczi E, Chindemi PA, Bolyos M. (1993) Am. J. Physiol. 264: G112 - G117.PubMedGoogle Scholar
  4. 4.
    McAbee, DD, Nowatzke W, Oehler C, Sitaram M, Sbaschnig E, Opferman JT, Carr J, Esbensen K. (1993) Biochemistry 32: 13749–13760.PubMedCrossRefGoogle Scholar
  5. 5.
    Ziere, GJ, Vandijk MCM, Bijsterbosch MK. Vanberkel TJC. (1992) J. Biol. Chem. 267: 11229–11235.Google Scholar
  6. 6.
    McAbee, DD, Esbensen K. (1991) J. Biol. Chem. 266: 23624–23631.Google Scholar
  7. 7.
    Schachter, H, Jabbal I, Hudgin RL, Pinteric L, McGuire EJ, Roseman S. (1970) J. Biol. Chem. 245: 1090–1100.Google Scholar
  8. 8.
    Ray, DA, Weigel PH. (1985) Anal. Biochem. 145: 37–46.PubMedCrossRefGoogle Scholar
  9. 9.
    Burnette, WN. (1981) Anal. Biochem. 112: 195–203.PubMedCrossRefGoogle Scholar
  10. Zeng, FY, Oka JA. Weigel PH. (1996) Glycobiology 6:247–255.Google Scholar
  11. 11.
    Leung, JO, Holland EC, Drickamer K. (1985) J. Biol. Chem. 260: 12523–12527.Google Scholar
  12. 12.
    Drickamer, K, Mamon JF, Bins G, Leung JO. (1984) J. Biol. Chem. 259: 770–778.Google Scholar
  13. 13.
    Ashwell, G, Harford J. (1982) Annu. Rev. Biochem. 51: 531–554.Google Scholar
  14. 14.
    Imber, MJ, Pizzo SV. (1983) Biochem. J. 212: 249–257.PubMedGoogle Scholar
  15. 15.
    Coddeville, B, Strecker G, Wieruszeski J-M, Vliegenthart JFG, van Halbeek H, Peter-Katalinic J, Egge H, Spik G. (1992) Carbohydrate Res. 236: 145–164.CrossRefGoogle Scholar
  16. 16.
    Spik, G, Coddeville B, Montreuil J. (1988) Biochimie 70: 1459–1469.PubMedCrossRefGoogle Scholar
  17. 17.
    Lce, YC, Townsend RR, Hardy MR, Lonngren J, Arnarp J, Haraldsson M, Lonn H. (1983) J. Biol. Chem. 258: 199–202.Google Scholar
  18. 18.
    Sitaram, MP, Maloney B, McAbee DD. (1996) FASEB J. 10: 1263a.Google Scholar
  19. 19.
    Moguilevsky, N, Retegui LA, Courtoy PJ, Castracane CE, Masson PL. (1984) Lab. Invest. 50: 335–340.PubMedGoogle Scholar
  20. 20.
    Herzig, MCS, Weigel PH. (1990) Biochemistry 29: 6437–6447.PubMedCrossRefGoogle Scholar
  21. 21.
    Sawyer, JT, Sanford JP, Doyle D. (1988) J. Biol. Chem. 263: 10534–10538.Google Scholar
  22. 22.
    Weigel, PH, Oka JA. (1983) J. Biol. Chem. 258: 5095–5102.Google Scholar
  23. 23.
    McAbee, DD, Ling YY. (1997) J. Cell. Physiol. 171: 75–86.Google Scholar
  24. 24.
    Kelm, S, Schauer R. (1988) Biol. Chem. Hoppe-Seyler 369: 693–704.Google Scholar
  25. 25.
    li, M, Kurata H, Itoh N, Yamashina I, Kawasaki T. (1990) J. Biol. Chem. 265: 11295–11298.Google Scholar
  26. 26.
    Ozaki, K. Ii M, Itoh N, Kawasaki T. (1992) J. Biol. Chem. 267: 9229–9235.Google Scholar
  27. 27.
    Britigan, BE, Ratcliffe HR, Buettner GR, Rosen GM. (1996) Biochim. Biophys. Acta 1290: 231–240.Google Scholar
  28. 28.
    Courtoy, PJ, Moguilevsky N, Retegui LA, Castracane CE, Masson PL. (1984) Lab. Invest. 50: 329–334.PubMedGoogle Scholar
  29. 29.
    Hoyle, GW, Hill RL. (1991) J. Biol. Chem. 266: 1850–1857.Google Scholar
  30. 30.
    Kempka, G, Roos PH, Kolb-Bachofen V. (1990) J. Immunol. 144: 1004–1009.PubMedGoogle Scholar
  31. 31.
    Ishibashi, S, Hammer RE, Herz J. (1994) J. Biol. Chem. 269: 27803–27806.Google Scholar

Copyright information

© Springer Science+Business Media New York 1998

Authors and Affiliations

  • Douglas D. McAbee
    • 1
  • David J. Bennatt
    • 1
  • Yuan Yuan Ling
    • 1
  1. 1.Department of Biological SciencesUniversity of Notre DameNotre DameIndiana

Personalised recommendations