Factor H co-purifies with thrombospondin isolated from platelet secretate

J. A. Carron, R. C. Bates, A. I. Smith, T. Tetoz, A. Arellano, D. L. Gordon, G. F. Burns

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12 Citations (Scopus)

Abstract

Thrombospondin is a trimeric glycoprotein that has several known functions, including roles in platelet aggregation, phagocytosis and an inhibitor of angiogenesis. Typically the molecule is isolated from platelet secretate by heparin affinity followed by sizing chromatography. In this study, purity is analysed by 7.5% SDS-PAGE under reducing conditions when thrombospondin monomers run as a band at around 180 kDa. Under nonreducing conditions of 7.5% SDS-PAGE, thrombospondin does not penetrate beyond the stacking gel; however, under these conditions a major contaminating band can be seen which, upon reduction, merges into the thrombospondin band. Further purification of this contaminating protein was achieved by DEAE chromatography and it was identified as Factor H by peptide sequencing and immunoblotting. Factor H function was demonstrated by the ability of the protein to function as a cofactor in die Factor-I-mediated cleavage of C3b. Since Factor H has several known functions, such contamination could confound functional studies of thrombospondin thus purified and a pre-elution step of the heparin affinity column is recommended.

Original languageEnglish
Pages (from-to)305-311
Number of pages7
JournalBiochimica et Biophysica Acta - General Subjects
Volume1289
Issue number3
DOIs
Publication statusPublished - 17 Apr 1996
Externally publishedYes

Keywords

  • Contamination
  • Factor H
  • Glycoprotein
  • Heparin affinity
  • Thrombospondin

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