Identification of immunoglobulin heavy chain binding protein as glucose-regulated protein 78 on the basis of amino acid sequence, immunological cross-reactivity, and functional activity

J Cell Sci Suppl. 1989;11:115-37. doi: 10.1242/jcs.1989.supplement_11.10.

Abstract

Immunoglobulin heavy chain binding protein (BiP) associates transiently with various proteins destined for the secretory pathway. To investigate the relationship between BiP and the 78K (K = 10(3) Mr) glucose-regulated protein (GRP78), we have determined a partial amino acid sequence of purified mouse BiP and isolated and sequenced a full-length cDNA clone encoding mouse GRP78. The 26 amino-terminal residues of the mature BiP protein are identical to a sequence of amino acids located near the start of the open reading frame encoding GRP78. A polyclonal antiserum raised against mouse GRP78 protein expressed in bacteria from the cloned GRP78 cDNA could immunoprecipitate complexes consisting of BiP and unfolded forms of immunoglobulin heavy chains. Furthermore, a monoclonal antibody raised against mouse BiP immunoprecipitated mouse GRP78 expressed in monkey CV-1 cells from an SV40-GRP78 recombinant vector. Finally, like the endogenous BiP of simian cells, mouse GRP78 associated with malfolded, non-glycosylated forms of influenza hemagglutinin (HA) when GRP78 and HA were co-expressed from SV40 vectors in CV-1 cells. These studies confirm that BiP is identical to GRP78. Comparison of the nucleic acid and deduced amino acid sequence of mouse GRP78 with those of other rodent and human GRP78s revealed an extremely high degree of sequence identity. BiP/GRP78 is closely related (approximately 60% identity) to the cytoplasmic 70K heat-shock proteins. Surprisingly, the carboxy-terminal 29 amino acids of BiP/GRP78, which are not conserved in HSP70 proteins, are almost identical in sequence to the steroidogenesis activator peptide found in the cytoplasm of rat Leydig tumor cells. Possible relationships between these polypeptides are discussed.

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
  • Carrier Proteins* / biosynthesis
  • Carrier Proteins* / immunology
  • Carrier Proteins* / isolation & purification
  • Cell Line
  • Chromatography, Agarose
  • Cloning, Molecular
  • Cross Reactions
  • DNA / genetics
  • Escherichia coli / metabolism
  • Fluorescent Antibody Technique
  • Genetic Vectors
  • Heat-Shock Proteins*
  • Immunoglobulin Heavy Chains*
  • Mice
  • Molecular Chaperones*
  • Molecular Sequence Data
  • Precipitin Tests
  • Recombinant Fusion Proteins / biosynthesis
  • Simian virus 40 / genetics

Substances

  • Carrier Proteins
  • Heat-Shock Proteins
  • Immunoglobulin Heavy Chains
  • Molecular Chaperones
  • Recombinant Fusion Proteins
  • DNA
  • molecular chaperone GRP78