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The Distal Hinge of the Reactive Site Loop and Its Proximity
The serpin plasminogen activator inhibitor type 1 (PAI-1) plays a regulatory role in various physiological processes ( e.g. fibrinolysis and pericellular proteolysis) and forms a potential target for therapeutic interventions. In this study we identified the epitopes of three PAI-1 inhibitory monocl...
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Published in: | The Journal of biological chemistry 2001-11, Vol.276 (48), p.44912-44918 |
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Main Authors: | , , , , , , , |
Format: | Article |
Language: | English |
Citations: | Items that this one cites Items that cite this one |
Online Access: | Get full text |
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Summary: | The serpin plasminogen activator inhibitor type 1 (PAI-1) plays a regulatory role in various physiological processes ( e.g. fibrinolysis and pericellular proteolysis) and forms a potential target for therapeutic interventions. In this study we identified
the epitopes of three PAI-1 inhibitory monoclonal antibodies (MA-44E4, MA-42A2F6, and MA-56A7C10). Differential cross-reactivities
of these monoclonals with PAI-1 from different species and sequence alignments between these PAI-1s, combined with the three-dimensional
structure, revealed several charged residues as possible candidates to contribute to the respective epitopes. The production,
characterization, and subsequent evaluation of a variety of alanine mutants using surface plasmon resonance revealed that
the residues His 185 , Arg 186 , and Arg 187 formed the major sites of interaction for MA-44E4. In contrast, the epitopes of MA-42A2F6 and MA-56A7C10 were found to be
conformational. The epitope of MA-42A2F6 comprises residues Lys 243 and Glu 350 , whereas the epitope of MA-56A7C10 comprises residues Glu 242 , Lys 243 , Glu 244 , Glu 350 , Asp 355 , and Arg 356 . The participation of Glu 350 , Asp 355 , and Arg 356 provides a molecular explanation for the differential exposure of this epitope in the different conformations of PAI-1 and
for the effect of these antibodies on the kinetics of the formation of the initial PAI-1-proteinase complexes. The localization
of the epitopes of MA-44E4, MA42A2F6, and MA-56A7C10 elucidates two previously unidentified molecular mechanisms to modulate
PAI-1 activity and opens new perspectives for the rational development of PAI-1 neutralizing compounds. |
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ISSN: | 0021-9258 1083-351X |
DOI: | 10.1074/jbc.M103077200 |