Expression, purification and characterization of recombinant murine granulocyte-macrophage colony-stimulating factor and bovine interleukin-2 from yeast

Gene. 1987;55(2-3):287-93. doi: 10.1016/0378-1119(87)90288-5.

Abstract

Expression and secretion of two lymphokines, murine granulocyte-macrophage colony-stimulating factor (MuGM-CSF) and bovine interleukin-2 (BoIL-2), to levels of 50-60 mg per liter were achieved by placing these cDNAs in a Saccharomyces cerevisiae expression vector that utilized the yeast alcohol dehydrogenase-2 promoter and alpha-factor leader peptide. These lymphokines were purified to homogeneity by direct application of the crude yeast medium to reversed-phase high-performance liquid chromatography. Despite the fact that both lymphokines contain at least one N-glycosylation site and have identical N-terminal residues (Ala-Pro-Thr), recombinant (R) GM-CSF was found to be heterogeneously glycosylated by yeast while RBoIL-2 was secreted without glycosylation. Additionally, approximately 40% of the RGM-CSF was found to be proteolytically cleaved after the second amino acid residue, while RBoIL-2 was found to be intact.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Cattle
  • Colony-Stimulating Factors / biosynthesis*
  • Colony-Stimulating Factors / isolation & purification
  • Colony-Stimulating Factors / metabolism
  • Granulocytes
  • Interleukin-2 / biosynthesis*
  • Interleukin-2 / isolation & purification
  • Interleukin-2 / metabolism
  • Macrophages
  • Mice
  • Promoter Regions, Genetic
  • Protein Processing, Post-Translational
  • Protein Sorting Signals
  • Recombinant Fusion Proteins / biosynthesis*
  • Recombinant Fusion Proteins / isolation & purification
  • Recombinant Fusion Proteins / metabolism
  • Recombinant Proteins / biosynthesis*
  • Saccharomyces cerevisiae / genetics
  • Saccharomyces cerevisiae / metabolism

Substances

  • Colony-Stimulating Factors
  • Interleukin-2
  • Protein Sorting Signals
  • Recombinant Fusion Proteins
  • Recombinant Proteins