Alternatively spliced isoform of P-selectin is present in vivo as a soluble molecule

Norihisa Ishiwata, Koji Takio, Masahiko Katayama, Kiyoaki Watanabe, Koiti Titani, Yasuo Ikeda, Makoto Handa

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108 Citations (Scopus)

Abstract

To demonstrate the presence of a soluble isoform of P-selectin predicted from cDNA sequencing (Johnston, G. I., Bliss, G. A., Newman, P. J., and McEver, R. P. (1990) J. Biol. Chem. 265, 21381-21385), we immunoisolated and compared structurally P-selectin from fresh frozen human plasma with that from washed intact platelets. Plasma P-selectin was reactive with rabbit antiserum to a synthesized peptide (residues 762-774 of mature P-selectin) but was significantly less reactive with antibody to a peptide (residues 747- 760). In contrast, platelet P-selectin reacted with both antibodies. S- Pyridylethylated plasma P-selectin was fractionated by reversed phase-high performance liquid chromatography into two major species. From platelets, two virtually identical species were separated. Sequential digestion with Achromobacter protease I and then Staphylococcus V8 protease produced peptides assigned to the tail region of the protein including the putative spliced site. From the more hydrophilic species in both plasma and platelets, a peptide completely lacking the sequence of the putative spliced site was identified. In contrast, the more hydrophobic species yielded a peptide with an intact transmembrane sequence. Hence, these results provide direct evidence that the previously predicted soluble isoform of P-selectin is actually synthesized in vivo and is present as a circulating molecule.

Original languageEnglish
Pages (from-to)23708-23715
Number of pages8
JournalJournal of Biological Chemistry
Volume269
Issue number38
Publication statusPublished - 1994 Sep 23

ASJC Scopus subject areas

  • Biochemistry
  • Molecular Biology
  • Cell Biology

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    Ishiwata, N., Takio, K., Katayama, M., Watanabe, K., Titani, K., Ikeda, Y., & Handa, M. (1994). Alternatively spliced isoform of P-selectin is present in vivo as a soluble molecule. Journal of Biological Chemistry, 269(38), 23708-23715.