Molecular heterogeneity of tumor rejection antigen/heat shock protein GP96.
Glycoproteins of 96 kDa (gp96) have been shown to mediate tumor-specific immunogenicity of a number of murine sarcomas. The purity of gp96 preparations used in these studies was originally demonstrated by their homogeneity on silver-stained gels and the observation that a single amino acid terminus was detected by micro-sequencing. Results reported here show that gp96 preparations consist of 3-4 closely spaced bands. However, each of the bands is recognized by well-characterized monoclonal or monospecific antibodies to gp96. We have obtained purified preparations of a slowly migrating 110-kDa and a faster migrating 96-kDa band of the gp96 cluster from the Meth A sarcoma and have observed both species to be immunogenic in a dose-restricted manner. In addition to the size-based heterogeneity, purified 96-kDa molecules are found to contain 2 major populations, which share the same amino and carboxy termini but differ in the region recognized by an anti-grp94 monoclonal antibody (MAb). The heterogeneity in gp96 preparations does not result from differences in glycosylation but may result from differential phosphorylation, conformation or the antigenic peptides associated with gp96. It has been suggested that the tumor-specific immunogenicity of gp96 preparations does not derive from gp96 but from a 110-kDa protein which co-purifies with gp96 and is distinct from it. Our observations show instead that the presence of a number of bands in purified gp96 preparations is due to heterogeneity within gp96 molecules rather than to contamination with unrelated proteins.
Feldweg
A M
AM
Department of Biological Sciences, Fordham University, Bronx, NY 10458, USA.
Srivastava
P K
PK
D000818:Animals; D000911:Antibodies, Monoclonal / Q000276:immunology; D000951:Antigens, Neoplasm / Q000737:chemistry* / Q000276:immunology; D015153:Blotting, Western; D018841:HSP90 Heat-Shock Proteins / Q000737:chemistry* / Q000276:immunology; D051379:Mice; D008807:Mice, Inbred BALB C; D008970:Molecular Weight
D000911:Antibodies, Monoclonal; D000951:Antigens, Neoplasm; D018841:HSP90 Heat-Shock Proteins; C052646:sarcoma glycoprotein gp96 rejection antigens