Summary
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 × 10−8)
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, complex-ing of tPA with Α1arantitrypsin 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.
