Type I and II collagens together with other extracellular matrix molecules are degraded in normal remodeling associated with physiological processes such as morphogenesis, growth, wound healing and physiological bone turnover. In addition, extracellular matrix is degraded during pathological processes, such as arthristis, osteolysis or metastasis. The goal of this research was to characterize HLA-DR1 and HLA-DR 4 restricted epitopes derived from the processing of collagen I and II by extracellular metalloproteases (MMPs), which could be different from the epitopes generated by the action of cathepsins, during the endocytic pathway of antigen processing. Complete sequences of peptides proteolyzed from highly purified human collagen-I and II were determined using both recombinant enzymes (MMPs and cathepsins) as well as purified dendritic cell (DC)-derived plasma membranes and endosomal fractions coupled with proteomics methods, such as de novo peptide sequencing using a nanospray LC-MS/MS on a LTQ linear ion trap mass spectrometer interfaced with a triversa nanomate nanoelectrospray ion source tandem mass spectroscopy (MS/MS). A wide variety of peptides originated from collagen-I and II were shown to have cleavage sites specific for metalloproteases and typical hydroxylations of proline and lysine, as posttranslational modifications. IC50 and binding affinities of some peptides epitopes derived from collagen II were characterized using recombinant HLA-DR1 and HLA-DR4 molecules and their immunogenicity was determined using transgenic mice for these proteins

Characterization of new peptide epitopes derived from collagen I and II processing by metalloproteases associated with human dendritic cells

241st National Meeting (2011)

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