Biochemical and cytological characterization of DROP-1: a widely distributed proteoglycan in Drosophila

Michael Graner, Kathy Stupka, Timothy L. Karr

Research output: Contribution to journalArticle

18 Scopus citations

Abstract

Using Drosophila testis as a source of antigen, 12 monoclonal antibodies were isolated that all recognize a set of three high molecular weight molecules present on Drosophila sperm and also in the fertilized egg. Among these antibodies, one is highly specific for sperm, while the remaining 11 detect epitopes present not only on sperm, but also in yolk spheres or in a punctate distribution in the egg. Here we cytologically and biochemically characterize the (common) antigens to five of these antibodies. Several biochemical properties suggest that these antibodies recognize a family of glycosaminoglycan-containing proteoglycans: (1) three diffuse, poorly focused high molecular weight bands, all in excess of 200,000 Da were observed on Western blots of denaturing SDS gels; (2) all three bands have a pI in the range of 3.0-3.5; (3) the molecules are strongly resistant to proteolysis; (4) mild periodate oxidation renders the molecules reactive towards the derivatizing agent digoxygenin-hydrazide, indicating the likely presence of saccharide moieties; (5) trifluoromethylsulfonic acid treatment, which removes saccharide moieties, shifts the pI to 7.0; (6) β-elimination increases electrophoretic mobility of the antigens on SDS gels; (7) nitrous acid treatment, which cleaves N-sulfated glycosaminoglycans, also increases the electrophoretic mobility of the antigens on SDS gels. We conclude that the antigens recognized by these antibodies are likely to be heparan sulfate proteoglycans. These results indicate that DROP-1 may represent a family of proteoglycans present during embryogenesis and later stages of development in Drosophila. DROP-1 represents the third proteoglycan to be characterized in Drosophila.

Original languageEnglish (US)
Pages (from-to)557-567
Number of pages11
JournalInsect Biochemistry and Molecular Biology
Volume24
Issue number6
DOIs
StatePublished - Jun 1994
Externally publishedYes

Keywords

  • Fertilization
  • Monoclonal antibodies
  • Proteoglycans
  • Sperm

ASJC Scopus subject areas

  • Biochemistry
  • Molecular Biology
  • Insect Science

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