Cloning of the cDNA for rabbit L-selectin and expression of recombinant protein with a kinase target site to facilitate radiolabeling

Biochem Biophys Res Commun. 1996 Aug 14;225(2):406-12. doi: 10.1006/bbrc.1996.1187.

Abstract

The cDNA encoding rabbit L-Selectin has been cloned from a cDNA library, utilizing a PCR-derived probe. It encodes a peptide of 377 amino acids, including a signal peptide of 38 amino acids. Sequence analysis demonstrated extensive homology with L-Selectin's from other species. Recombinant rabbit L-Selectin protein was expressed in eukaryotic cells in a chimeric construct incorporating the entire extracellular portion of the protein coding region, a phosphokinase target site to allow high activity radiolabeling with 32P, and the constant region of the heavy chain of human IgG1 to facilitate purification and detection. Amino-terminal peptide sequencing of recombinant L-Selectin confirmed that the signal peptide had been removed at the expected site.

Publication types

  • Research Support, Non-U.S. Gov't
  • Research Support, U.S. Gov't, P.H.S.

MeSH terms

  • Amino Acid Sequence
  • Animals
  • Base Sequence
  • Cell Line
  • Cloning, Molecular
  • DNA, Complementary
  • Humans
  • L-Selectin / genetics*
  • L-Selectin / metabolism
  • Molecular Sequence Data
  • Phosphorus Radioisotopes
  • Phosphotransferases / metabolism
  • Rabbits
  • Recombinant Proteins / genetics
  • Recombinant Proteins / metabolism
  • Sequence Homology, Amino Acid

Substances

  • DNA, Complementary
  • Phosphorus Radioisotopes
  • Recombinant Proteins
  • L-Selectin
  • Phosphotransferases