SLA/LP/tRNP((Ser)Sec) antigen in autoimmune hepatitis: identification of the native protein in human hepatic cell extract

J Autoimmun. 2010 Feb;34(1):59-65. doi: 10.1016/j.jaut.2009.07.005. Epub 2009 Aug 14.

Abstract

A diagnostic subgroup of AIH type 1 is characterized by specific serum antibodies against soluble liver protein. The respective autoantigen was named SLA/LP/tRNP((Ser)Sec), after three homologous recombinant polypeptides were isolated from expression gene libraries. We analyzed human cultured liver cells for the human homologue of recombinant SLA/LP/tRNP((Ser)Sec) by antigen purification. In addition, a monoclonal antibody was generated against recombinant SLA-p35, a truncated recombinant SLA-reactive polypeptide. With a positive patient serum, immune affinity chromatography was performed on the 52 kD-SLA main antigenic determinant pre-enriched by ion exchange chromatography. By mass spectrometry, the 52 kD-SLA/LP/tRNP ((Ser)Sec) autoantigen was unambiguously identified in the purification product. The identity of the recombinant SLA-p35 and its human homologue was further confirmed by a specific signal of the anti SLA-p35 monoclonal antibody with purified human SLA/LP/tRNP((Ser)Sec). The 48 kD-SLA species frequently comigrating in SLA-immunoblotting however was not identified by either approach. We conclude that the native counterpart of recombinant tRNP((Ser)(Sec)) indeed is detectable with a molecular weight of 52 kD in soluble liver extract of human cells as the major antigenic component of SLA/LP/tRNP((Ser)Sec).

MeSH terms

  • Amino Acid Sequence
  • Antibodies, Monoclonal / metabolism*
  • Autoantigens / blood
  • Autoantigens / genetics
  • Autoantigens / immunology
  • Autoantigens / metabolism*
  • Cell Extracts
  • Cell Line, Tumor
  • Chromatography, Affinity
  • Cloning, Molecular
  • Hepatitis, Autoimmune / blood
  • Hepatitis, Autoimmune / diagnosis
  • Hepatitis, Autoimmune / immunology
  • Hepatitis, Autoimmune / metabolism*
  • Hepatocytes / immunology
  • Hepatocytes / metabolism*
  • Hepatocytes / pathology
  • Humans
  • Mass Spectrometry
  • Molecular Sequence Data
  • Peptide Fragments / blood
  • Peptide Fragments / genetics
  • Peptide Fragments / immunology
  • Peptide Fragments / metabolism
  • RNA, Transfer, Amino Acyl / blood
  • RNA, Transfer, Amino Acyl / genetics
  • RNA, Transfer, Amino Acyl / immunology
  • RNA, Transfer, Amino Acyl / metabolism*
  • Recombinant Proteins / blood
  • Recombinant Proteins / genetics
  • Recombinant Proteins / immunology
  • Recombinant Proteins / metabolism

Substances

  • Antibodies, Monoclonal
  • Autoantigens
  • Cell Extracts
  • Peptide Fragments
  • RNA, Transfer, Amino Acyl
  • Recombinant Proteins
  • selenocysteinyl-tRNA