title: The conformation changes of the finger domain of tissue type plasminogen activator during the activator-inhibitor reaction.
abstract: A peptide fragment of tissue plasminogen activator (tPA) corresponding to amino acid residues 4-8 (tPA4-8) was synthesized, coupled to thyroglobulin and injected into rabbits. Antibodies specific to the peptide tPA4-8 were purified immunochemically on the pentapeptide coupled to CNBr-Sepha rose 4B. Anti-tPA4-8 antibodies, reacted with iodinated peptide tPA4-8, showing a relatively high binding affinity (KD = 2.3 x 10(-8) M). There was no interaction between anti-tPA4-8 antibodies and native one- or two-chain tPA. However, reduction of disulfide bonds unmasked the epitope on the heavy chain of tPA which became accessible to anti-tPA4-8 antibodies. Similarly, complexing of tPA with alpha 1-antitrypsin inhibitor resulted in unmasking of the epitope formed by amino acid residues in the positions 4-8. Presented data suggest that complexing of tPA with inhibitors results in conformational changes occurring in the "finger" domain of tPA molecule and such conformational transition can be detected by antipeptide antibodies. Therefore, anti-tPA4-8 antibodies may be employed as sequence-specific reporter molecules to monitor local conformational changes in tPA molecule.
authors.affiliation: Department of Biophysics, Institute of Physiology and Biochemistry, Medical School, Lódź, Poland.
authors.forename: M
authors.initials: M
authors.lastname: Wilczyńska
authors.forename: C S
authors.initials: CS
authors.lastname: Cierniewski
mesh_terms: D000595:Amino Acid Sequence; D015153:Blotting, Western; D007120:Immunochemistry; D008969:Molecular Sequence Data; D010446:Peptide Fragments / Q000138:chemical synthesis / Q000276:immunology; D015849:Plasminogen Inactivators / Q000378:metabolism*; D011485:Protein Binding; D011487:Protein Conformation; D010959:Tissue Plasminogen Activator / Q000276:immunology / Q000378:metabolism; D000515:alpha 1-Antitrypsin / Q000378:metabolism
chemical_list: D010446:Peptide Fragments; D015849:Plasminogen Inactivators; D000515:alpha 1-Antitrypsin; D010959:Tissue Plasminogen Activator