Identification of a Target Site in Plasminogen Activator Inhibitor-1 That Allows Neutralization of Its Inhibitory Properties Concomitant with an Allosteric Up-regulation of Its Antiadhesive Properties

Ngo, Thu-Hoa; Hoylaerts, Marc; Knockaert, I; Brouwers, Els; Declerck, Paul

doi:10.1074/jbc.m008241200

Cited by 24 publications

(18 citation statements)

References 37 publications

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“…Conversion of PAI-1 between its active and latent forms is regulated by vitronectin, which circulates in plasma but is also a major constituent of the extracellular matrix [28,29]. In the data adjusted for total protein levels, an increase in activity without a change in antigen levels was still observed, suggesting that PAI activity is dependent on protein/vitronectin influx and subsequent stabilisation of PAI activity during high pressureamplitude ventilation.…”

Section: Discussionmentioning

confidence: 80%

Mechanical ventilation affects alveolar fibrinolysis in LPS-induced lung injury

Dahlem¹,

Bos²,

Haitsma³

et al. 2006

Eur Respir J

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The aim of the present study was to determine the effects of mechanical ventilation on alveolar fibrin turnover in lipopolysaccharide (LPS)-induced lung injury.In a randomised controlled trial, Sprague-Dawley rats (n561) were allocated to three ventilation groups after intratracheal LPS (Salmonella enteritidis) instillations. Group I animals were subjected to 16 cmH 2 O positive inspiratory pressure (PIP) and 5 cmH 2 O positive end-expiratory pressure (PEEP); group II animals to 26 cmH 2 O PIP and 5 cmH 2 O PEEP; and group III animals to 35 cmH 2 O PIP and 5 cmH 2 O PEEP. Control rats (not mechanically ventilated) received LPS. Healthy rats served as a reference group. Levels of thrombin-antithrombin complex (TATc), D-dimer, plasminogen activator inhibitor (PAI) activity and PAI-1 antigen in bronchoalveolar lavage fluid were measured.LPS-induced lung injury increased TATc, D-dimer and PAI activity and PAI-1 antigen levels versus healthy animals. High pressure-amplitude ventilation increased TATc concentrations.D-dimer concentrations were not significantly raised. Instead, PAI activity increased with the amplitude of the pressure, from 0.7 U?mL -1 in group I to 3.4 U?mL -1 in group II and 5.0 U?mL -1 in group III. There was no change in PAI-1 antigen levels.In conclusion, mechanical ventilation creates an alveolar/pulmonary anti-fibrinolytic milieu in endotoxin-induced lung injury which, at least in part, might be due to an increase in plasminogen activator inhibitor activity.

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Section: Discussionmentioning

confidence: 80%

Mechanical ventilation affects alveolar fibrinolysis in LPS-induced lung injury

Dahlem¹,

Bos²,

Haitsma³

et al. 2006

Eur Respir J

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“…Some PAI-1 variants with single mutations and modest decreases in the rate of latency transition have been obtained through heuristic protein engineering (78,79) while others have been identified by chance (80)(81)(82)(83). The variants with the slowest latency transition carry multiple mutations and have been obtained through in vitro molecular evolution, i.e.…”

Section: Pai-1 Latency Transitionmentioning

confidence: 99%

Biochemical properties of plasminogen activator inhibitor-1

Dupont

Madsen²,

Kristensen³

et al. 2009

Front Biosci

102

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PAI-1 is a Mr ~50,000 glycoprotein, which is the primary physiological inhibitor of the two plasminogen activators uPA and tPA. PAI-1 belongs to the serpin protein family. Studies of PAI-1 have contributed significantly to the elucidation of the protease inhibitory mechanism of serpins, which is based on a metastable native state becoming stabilised by insertion of the RCL into the central beta-sheet A and formation of covalent complexes with target proteases. In PAI-1, this insertion can occur in the absence of the protease, resulting in generation of a so-called latent, inactive form of the protein. PAI-1, in its active state, also binds to the extracellular protein vitronectin. When in complex with its target proteases, it binds with high affinity to endocytosis receptors of the low density receptor family.

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“…The epitope of CB5B10 was localized in ␣-helix E and the turn connecting helix E and strand s1A (52). Epitope mapping of the inhibitory anti-rat PAI-1 MA-124K1 revealed the major contribution of residues Glu 212 and Glu 220 localized on strand s1B and s2B (53). For the monoclonal antibodies I-201 (noninhibitory) and M-5 (inhibitory), the dominant residues were found to be Gln 56 (located in helix C) and Asp 181 (ts3As4C), respectively (49).…”

Section: Discussionmentioning

confidence: 98%

The Distal Hinge of the Reactive Site Loop and Its Proximity

Bijnens¹,

Gils²,

Stassen³

et al. 2001

Journal of Biological Chemistry

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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 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.Plasminogen activator inhibitor type 1 (PAI-1), 1 a member of the serpin (serine proteinase inhibitor) superfamily (1-4), controls the plasminogen system at the level of tissue-type and urokinase-type plasminogen activator (t-PA and u-PA, respectively). Because PAI-1 is the main physiological inhibitor of t-PA in plasma (5), increased levels of PAI-1 result in a hypofibrinolytic state and are correlated with various vascular disorders such as venous thrombo-embolism, coronary artery disease, myocardial infarction, and atherosclerosis (6 -9). The u-PA-inhibiting effect of PAI-1 has its main physiological implications in processes outside of the circulation (10, 11).PAI-1 is a unique serpin because of its functional and conformational flexibility (12). PAI-1, synthesized as an active molecule, converts spontaneously into a nonreactive, latent form, which can be partially reactivated by denaturing reagents (13). Additionally, a third distinct, noninhibitory form (substrate) is identified that is reactive toward its target proteinases without the formation of a stable complex (14 -16).Elucidation of the three-dimensional structure of active PAI-1 (17, 18) reveals that the N-terminal side of the reactive site loop (extended from P16 to P3Ј and including the bait peptide bond Arg 345 -Met 346 (P1-P1Ј)) is exposed and accessible for the target proteinase. The C-terminal side of the reactive site loop (P4Ј-P13Ј) forms strand s1C in ␤-sheet C.Conversion to the latent state implies the insertion of the N-terminal side of the reactive site loop into ␤-sheet A, the loss of strand s1C from ␤-sheet C, and the formation of an unusual extended loop by the C-terminal side of the reactive site loop, resulting in the distortion of the P1-P1Ј "bait" peptide bond (19). It is hypothesized that th...

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Identification of a Target Site in Plasminogen Activator Inhibitor-1 That Allows Neutralization of Its Inhibitory Properties Concomitant with an Allosteric Up-regulation of Its Antiadhesive Properties

Cited by 24 publications

References 37 publications

Mechanical ventilation affects alveolar fibrinolysis in LPS-induced lung injury

Mechanical ventilation affects alveolar fibrinolysis in LPS-induced lung injury

Biochemical properties of plasminogen activator inhibitor-1

The Distal Hinge of the Reactive Site Loop and Its Proximity

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