title: Analysis of the extracellular processing of HIV-1 gp160-derived peptides using monoclonal antibodies specific to H-2Dd molecule complexed with p18-I10 peptide.
abstract: The immunodominant peptide of human immunodeficiency virus 1 gp 160 for murine cytotoxic T cells of H-2d haplotype, has been originally identified as a 15 amino acid residue peptide P18IIIB (RIQRGPGRAFVTIGK) (Takahashi et al., 1988). Further studies have indicated that a more active form of the peptide is generated by removal of the C-terminal dipeptide by angiotensin-I-converting enzyme (ACE), and additional detailed studies have shown that the actual immunodominant peptide is a decamer P18-I10 (RGPGRAFVTI) (Kozlowski et al., 1993). The effect of proteolytic processing on the antigenicity of P18IIIB peptide and its analogs was investigated by functional T cell assays based on the ability of T cell receptor (TCR) to recognize a specific major histocompatibility complex class I (MHC-I)/peptide complex. Recently we described a new monoclonal antibody (MAb) KP15 directed against the MHC-I molecule H-2Dd complexed with the 10-mer peptide P18-I10. Using this MAb, the cell surface H-2Dd/P18-I10 complex can be easily detected by flow cytometry (Polakova et al., 2000). Here we examined whether peptides longer than P18-I10 decamer form H-2Dd complexes recognized by KP15 MAb. Further we also analyzed how the ACE processing of P18IIIB-related peptides of different length affects their ability to form complexes with H-2Dd recognized by MAb KP15. These experiments confirmed that the ACE digestion of 15-mer peptide P18IIIB is the most effective in the production of a peptide capable of forming complex with H-2Dd recognized by KP15 MAb. The ACE digestion of longer peptides (16-mer to 19-mer) did not produce a significant quantity of peptides, capable of forming H-2Dd complexes recognizable with by MAb KP15. Peptides shorter than P18IIIB (13-mer to 10-mer), notably the optimally sized P18-I10 peptide lost most of their capacity to form H-2Dd complexes recognized by KP15 MAb. Our results show that the extracellular processing of MHC-I-restricted peptides, which cannot be overlooked in designing peptide-based vaccines, can be also studied by as simple and rapid assay as flow cytometry, provided MAbs specific to a particular MHC-I/peptide complex are available.
authors.affiliation: Cancer Research Institute, Slovak Academy of Sciences, Bratislava, Slovak Republic. exonpola@savba.sk
authors.forename: K
authors.initials: K
authors.lastname: Poláková
authors.forename: G
authors.initials: G
authors.lastname: Russ
mesh_terms: D000818:Animals; D000911:Antibodies, Monoclonal; D002460:Cell Line; D005434:Flow Cytometry; D006183:H-2 Antigens / Q000737:chemistry / Q000276:immunology; D019302:HIV Envelope Protein gp160 / Q000737:chemistry*; D015497:HIV-1 / Q000737:chemistry*; D051379:Mice; D010455:Peptides / Q000032:analysis* / Q000737:chemistry; D007703:Peptidyl-Dipeptidase A
chemical_list: D000911:Antibodies, Monoclonal; D006183:H-2 Antigens; D019302:HIV Envelope Protein gp160; D010455:Peptides; D007703:Peptidyl-Dipeptidase A