Isolation of ERp72 from guinea pig term placentae using heparin Sepharose affinity chromatography

Biochem Mol Biol Int. 1995 Aug;36(6):1143-52.

Abstract

The mammalian placenta synthesises many varied antigens, including proteins, such as hormones, enzymes and protease inhibitors. In this report, we isolated and purified the two protein isomerase-related protein precursor ERp72 isoforms from aqueous extracts of guinea pig placenta, by four (4) chromatographic procedures; i) affinity chromatography on immobilised heparin, ii) gel filtration (Ultrogel AcA-54), iii) anion exchange chromatography (Mono-Q), and, iv) negative immunoaffinity chromatography. From 20 term placentae, the final yield of ERp72 isoforms was 2.4mg (Mr 71.5 kDa) and 1.5mg (Mr 75.8 kDa). Identity was confirmed by NH2-terminal amino acid sequencing which demonstrated 85% homology to human ERp72. By indirect immunofluorecence. ER p72 expression was demonstrated in tunicamycin stressed pre-implantation embryos and unfertilised oocytes. These findings demonstrate the potential for immunological monitoring of ERp72 expression, by cultured oocytes and embryos, during manipulation by assisted reproductive technologies.

Publication types

  • Comparative Study
  • Research Support, Non-U.S. Gov't

MeSH terms

  • Amino Acid Sequence
  • Animals
  • Blastocyst / cytology
  • Blastocyst / metabolism
  • Chromatography, Affinity
  • Chromatography, Gel
  • Chromatography, Ion Exchange
  • Electrophoresis, Polyacrylamide Gel
  • Female
  • Fluorescent Antibody Technique
  • Guinea Pigs
  • Heparin
  • Humans
  • Membrane Glycoproteins / chemistry*
  • Membrane Glycoproteins / isolation & purification*
  • Mice
  • Molecular Sequence Data
  • Molecular Weight
  • Placenta / metabolism*
  • Pregnancy
  • Sepharose
  • Sequence Homology, Amino Acid

Substances

  • Membrane Glycoproteins
  • endoplasmic reticulum glycoprotein p72
  • Heparin
  • Sepharose