Plasmin generation induces neutrophil aggregation: dependence on the catalytic and lysine binding sites.

We established that plasmin (10(-10) M to 10(-6) M) caused neutrophils (PMN) to aggregate using an in vitro assay. Plasminogen had no PMN aggregatory activity even at a concentration of 2 microM. However, plasminogen caused PMN to aggregate when incubated with plasminogen activators [tissue plasminogen activator (25-200 U/ml) or urokinase (5-500 U/ml)]. Tissue plasminogen activator and urokinase alone had no PMN aggregatory activity. Analysis of these incubation mixtures indicated that plasmin was generated in the process and that the time course of plasmin generation correlated with the aggregation response. Active-site-inhibited plasmin did not induce PMN aggregation, indicating that a functional catalytic site was required for the response. Pretreatment of PMN with either active-site-inhibited plasmin or tranexamic acid prevented PMN aggregation by plasmin, indicating that both binding of plasmin to the cell surface via the lysine binding sites and catalysis were required for the response. The generation of plasmin during activation of fibrinolysis may play a pro-inflammatory role by mediating aggregation of PMN.     

Haemophilus influenzae uses the surface protein E to acquire human plasminogen and to evade innate immunity

Pathogenic microbes acquire the human plasma protein plasminogen to their surface. In this article, we characterize binding of this important coagulation regulator to the respiratory pathogen nontypeable Haemophilus influenzae and identify the Haemophilus surface protein E (PE) as a new plasminogen-binding protein. Plasminogen binds dose dependently to intact bacteria and to purified PE. The plasminogen-PE interaction is mediated by lysine residues and is also affected by ionic strength. The H. influenzae PE knockout strain (nontypeable H. influenzae 3655Δpe) bound plasminogen with ～65% lower intensity as compared with the wild-type, PE-expressing strain. In addition, PE expressed ectopically on the surface of Escherichia coli also bound plasminogen. Plasminogen, either attached to intact H. influenzae or bound to PE, was accessible for urokinase plasminogen activator. The converted active plasmin cleaved the synthetic substrate S-2251, and the natural substrates fibrinogen and C3b. Using synthetic peptides that cover the complete sequence of the PE protein, the major plasminogen-binding region was localized to a linear 28-aa-long N-terminal peptide, which represents aa 41-68. PE binds plasminogen and also vitronectin, and the two human plasma proteins compete for PE binding. Thus, PE is a major plasminogen-binding protein of the Gram-negative bacterium H. influenzae, and when converted to plasmin, PE-bound plasmin aids in immune evasion and contributes to bacterial virulence.     

Leishmania mexicana: LACK (Leishmania homolog of receptors for activated C-kinase) is a plasminogen binding protein

Leishmania mexicana is able to interact with the fibrinolytic system through its component plasminogen, the zymogenic form of the protease plasmin. In this study a new plasminogen binding protein of this parasite was identified: LACK, the Leishmania homolog of receptors for activated C-kinase. Plasminogen binds recombinant LACK with a K_d value of 1.6 ± 0.4 μM, and binding is lysine-dependent since it is inhibited by the lysine analog ε-aminocaproic acid. Inhibition studies with specific peptides and plasminogen binding activity of a mutated recombinant LACK have highlighted the internal motif ₂₆₀VYDLESKAV₂₆₈, similar to those found in several enolases, as involved in plasminogen binding. Recombinant LACK and secreted proteins, in medium conditioned by parasites, enhance plasminogen activation to plasmin by the tissue plasminogen activator (t-PA). In addition to its localization in the cytosol, in the microsomal fraction and as secreted protein in conditioned medium, LACK was also localized on the external surface of the membrane. The results presented here suggest that LACK might bind and enhance plasminogen activation in vivo promoting the formation of plasmin. Plasminogen binding of LACK represents a new function for this protein and might contribute to the invasiveness of the parasite.     

Identification of a novel plasmin(ogen)-binding motif in surface displayed alpha-enolase of Streptococcus pneumoniae

The interaction of Streptococcus pneumoniae with human plasmin(ogen) represents a mechanism to enhance bacterial virulence by capturing surface-associated proteolytic activity in the infected host. Plasminogen binds to surface displayed pneumococcal alpha-enolase (Eno) and is subsequently activated to the serine protease plasmin by host-derived tissue plasminogen activator (tPA) or urokinase (uPA). The C-terminal lysyl residues of Eno at position 433 and 434 were identified as a binding site for the kringle motifs of plasmin(ogen) which contain lysine binding sites. In this report we have identified a novel internal plamin(ogen)-binding site of Eno by investigating the protein-protein interaction. Plasmin(ogen)-binding activity of C-terminal mutated Eno proteins used in binding assays as well as surface plasmon resonance studies suggested that an additional binding motif of Eno is involved in the Eno-plasmin(ogen) complex formation. The analysis of spot synthesized synthetic peptides representing Eno sequences identified a peptide of nine amino acids located between amino acids 248-256 as the minimal second binding epitope mediating binding of plasminogen to Eno. Binding of radiolabelled plasminogen to viable pneumococci was competitively inhibited by a synthetic peptide FYDKERKVYD representing the novel internal plasmin(ogen)-binding motif of Eno. In contrast, a synthetic peptide with amino acid substitutions at critical positions in the internal binding motif identified by systematic mutational analysis did not inhibit binding of plasminogen to pneumococci. Pneumococcal mutants expressing alpha-enolase with amino acid substitutions in the internal binding motif showed a substantially reduced plasminogen-binding activity. The virulence of these mutants was also attenuated in a mouse model of intranasal infection indicating the significance of the novel plasminogen-binding motif in the pathogenesis of pneumococcal diseases.     

The fate of integrated DNA in Gaeumannomyces graminis transformants

Abstract The interaction of plasminogen with flagella of Escherichia coli was investigated. Plasminogen bound to flagella purified from E. coli LE392, a commonly used cloning host, and E. coli IH3069, and O25H1 strain isolated from a case of newborn bacteremia. The binding was inhibited by the lysine analog ϵ-aminocaproic acid, suggesting involvement of the lysine-binding Kringle domains of plasminogen in the binding. Purified flagella enhanced the formation of plasmin activity in the presence of tissue-type plasminogen activator; a similar enhancement was observed with flagella-expressing LE392 cells.     

Binding and activation of plasminogen on the platelet surface

A mechanism by which platelets might participate in fibrinolysis by binding plasminogen and influencing its activation has been examined. Binding of radioiodinated human Glu-plasminogen to washed human platelets was time-dependent and was enhanced 3-9-fold by stimulation of platelets with thrombin but not with ADP. The interaction with both stimulated and unstimulated cells was specific, saturable, divalent ion-independent, and reversible. The platelet-bound ligand had the molecular weight of plasminogen, and no conversion to plasmin was detected. Scatchard analyses provided evidence for a single class of plasminogen-binding sites on both stimulated and unstimulated cells. The Kd for thrombin-stimulated platelets was 2.6 +/- 1.3 microM, and 190,000 +/- 45,000 molecules were bound per cell, whereas unstimulated platelets bound 37,000 +/- 10,500 molecules/cell with a Kd of 1.9 +/- 0.15 microM. Plasminogen binding was inhibited in a dose-dependent manner by omega-aminocarboxylic acids at concentrations consistent with a requirement for an unoccupied high affinity lysine-binding site for plasminogen binding to the cells. When platelet-bound plasminogen was incubated with tissue plasminogen activator, urokinase, or streptokinase, gel analysis established that plasmin was preferentially associated with the platelet relative to the supernatant. Plasminogen and plasmin interacted with thrombin-stimulated platelets with similar binding characteristics, and there was no evidence for a binding site for plasmin which did not also bind plasminogen. Therefore, the results suggest that plasminogen activation is enhanced on the cell surface. In sum, these results indicate that platelets bind plasminogen at physiologic zymogen concentrations and this interaction may serve to localize and promote plasminogen activation.     

Fibronectin decreases the stimulatory effect of fibrin and fibrinogen fragment FCB-2 on plasmin formation by tissue plasminogen activator.

Fibronectin is a dimeric glycoprotein (Mr 440,000) involved in many adhesive processes. During blood coagulation it is bound and cross-linked to fibrin. Fibrin binding is achieved by structures (type I repeats) which are homologous to the "finger" domain of tissue plasminogen activator. Tissue plasminogen activator also binds to fibrin via the finger domain and additionally via the "kringle 2" domain. Fibrin binding of tissue plasminogen activator results in stimulation of its activity and plays a crucial role in fibrinolysis. Since fibronectin might interfere with this binding, we studied the effect of fibronectin on plasmin formation by tissue plasminogen activator. In the absence of fibrin, fibronectin had no effect on plasminogen activation. In the presence of stimulating fibrinogen fragment FCB-2, fibronectin increased the duration of the initial lag phase (= time period until maximally stimulated plasmin formation occurs) and decreased the rate of maximal plasmin formation which occurs after that lag phase mainly by increasing the Michaelis constant (Km). These effects of fibronectin were dose-dependent and were similar with single- and two-chain tissue plasminogen activator. They were also observed with plasmin-pretreated FCB-2. An apparent Ki of 43 micrograms/ml was calculated for the inhibitory effect of fibronectin when plasminogen activation by recombinant single-chain tissue plasminogen activator was studied in the presence of 91 micrograms/ml FCB-2. When a recombinant tissue plasminogen activator mutant lacking the finger domain was used in a system containing FCB-2, no effect of fibronectin was seen, indicating that the inhibitory effect of fibronectin might in fact be due to competition of fibronectin and tissue plasminogen activator for binding to fibrin(ogen) via the finger domain.     

Follicular plasminogen and plasminogen activator and the effect of plasmin on ovarian follicle wall.

Plasminogen, plasminogen activator, protease inhibitors, and a proteolytic activity are shown to be present in bovine follicular fluid. Much of the proteolytic activity appears to be due to plasmin. In addition, plasminogen activator activity can be demonstrated in follicle wall homogenates. Evidence that plasmin decreases the tensile strength of follicle wall preparations is also reported. The potential for the involvement of these substances in ovulation is discussed.     

Binding of vitronectin and plasminogen to Helicobacter pylori

Abstract We have studied how some extracellular matrix proteins, fibronectin, fibrinogen, collagen type I and type IV, plasminogen and vitronectin bind to Helicobacter pylori . Radiolabelled vitronectin and plasminogen bound to the haemagglutinating H. pylori strain 17874 at a high level (53% and 32%, respectively), type IV collagen showed an intermediate level of binding (16%), while binding by ¹²⁵I-labelled fibrinogen, fibronectin and collagen type I remained at a low level (5–7%). Both ¹²⁵I-vitronectin and plasminogen showed a dose-dependent binding to cells of H. pylori 17874. Plasminogen binding by this strain was specific since the binding was inhibited by nonlabelled plasminogen, but not by highly glycosylated glycoproteins such as fetuin and orosomucoid or by a variety of monosaccharides. We have previously shown that ¹²⁵I-vitronectin shows a specific and saturable binding to H. pylori 17874, and that sialic acid-rich glycoproteins such as fetuin and orosomucoid drastically reduced binding. We now report that a simultaneous incubation of ¹²⁵I-vitronectin and ¹²⁵I-plasminogen with cells of H. pylori 17874 showed a total binding approximately similar to the level of binding when either ¹²⁵I-plasminogen, or ¹²⁵I-vitronectin only were incubated with the bacterial cells. Nonlabelled vitronectin inhibited the binding of ¹²⁵I-plasminogen by H. pylori , but nonlabelled plasminogen had no effect on the binding of ¹²⁵I-vitronectin. Our findings suggest that there are different but probably closely localized binding sites for vitronectin and plasminogen on H. pylori 17874.     

Binding and activation of plasminogen on immobilized immunoglobulin G. Identification of the plasmin-derived Fab as the plasminogen-binding fragment

We have found that tissue plasminogen activator catalyzes the binding of plasminogen (Pg) to immunoglobulin G (IgG) immobilized on a surface. This enhancement is due to the formation of plasmin, since plasmin treatment of immobilized IgG produced a 20-fold increase in Pg binding. Pg binding is lysine site dependent and reversible. The augmentation of Pg binding by plasmin is specific as other proteases produced significantly less or no effect. Immobilized plasmin-treated IgG also specifically binds Pg in plasma. IgG-immobilized Pg is activated by tissue plasminogen activator, and a significant portion of the plasmin formed remains bound to the IgG. The Pg reactive species in a plasmin-treated IgG digest was identified as the Fab fragment by chromatography utilizing the immobilized high affinity lysine-binding site of plasminogen. Specificity of the interaction was further demonstrated by immunoblot-ligand analysis which demonstrated that the plasmin-derived Fab fragment bound Pg whereas papain-derived Fab or plasmin-derived Fc fragments did not. These data suggest that Pg binds to the new COOH-terminal lysine residue of the plasmin-derived Fab. Pg also binds to an immobilized immune complex following plasmin treatment. These findings indicate that surface-bound IgG localizes plasminogen thus extending the spectrum of activity of the plasmin system to immunologic reactions.     

The effects of plasminogen on in vitro ovine embryo development

Plasminogen activator production by ovine embryos and the effects of plasminogen on ovine embryo development and zona pellucida integrity were evaluated. Eight-cell to sixteen-cell embryos were cultured in Whitten's medium containing 0, 60, or 120 micrograms/ml plasminogen. Plasmin and plasminogen activator concentrations in the medium were determined by a caseinolytic assay. More blastocysts hatched in medium containing 60 and 120 micrograms/ml plasminogen (33 and 21%, respectively) than 0 microgram/ml plasminogen (0%; p less than 0.05). Zona pellucida dissolution time in acidified phosphate-buffered saline was less after incubation in medium with 60 and 120 micrograms/ml plasminogen (7.2 and 5.9 min, respectively) than 0 microgram/ml plasminogen (9.4 min; p less than 0.05). Plasminogen activator production was low until the morula stage, increased during morula-blastocyst transition, and remained elevated through blastocoelic expansion and hatching. Zona pellucida solubility, plasminogen activator production, and plasminogen conversion to plasmin increased as embryonic stage advanced; however, plasminogen activator production and plasmin conversion to plasmin were poorly correlated with zona pellucida solubility. The results indicate that ovine embryos produce plasminogen activator, and plasmin can increase zona pellucida solubility; however, other factors may also be involved in altering zona pellucida integrity prior to hatching.     

Effect of endogenous fibrinolysis activation on human plasminogen

Plasminogen preparation from donor blood and fibrinolytically active blood plasma from humans after sudden death were obtained using affinity chromatography on Lysin-sepharose 4B. The plasminogen preparation from donor blood was shown to be highly purified native plasminogen (Glu-plasminogen). The preparation containing activated plasminogen (Lys-plasminogen), plasmin, plasminogen activator, alpha 2-macroglobulin, alpha 1-antitrypsin, fibrin/fibrinogen was obtained from the blood plasma of humans after sudden death. The appearance of proteins lacking biological specificity to lysin-sepharose in the plasminogen preparation shows the ability of activated plasminogen and plasmin to form complexes with these proteins and demonstrates the retention of the functional activity in lysin-binding regions on their molecules. Monospecific sera to the isolated preparations were obtained, demonstrating the presence of the same immunochemical determinants in native and activated plasminogen.     

The plasminogen system and cell surfaces: evidence for plasminogen and urokinase receptors on the same cell type

The capacity of cells to interact with the plasminogen activator, urokinase, and the zymogen, plasminogen, was assessed using the promyeloid leukemic U937 cell line and the diploid fetal lung GM1380 fibroblast cell line. Urokinase bound to both cell lines in a time- dependent, specific, and saturable manner (Kd = 0.8-2.0 nM). An active catalytic site was not required for urokinase binding to the cells, and 55,000-mol-wt urokinase was selectively recognized. Plasminogen also bound to the two cell lines in a specific and saturable manner. This interaction occurred with a Kd of 0.8-0.9 microM and was of very high capacity (1.6-3.1 X 10(7) molecules bound/cell). The interaction of plasminogen with both cell types was partially sensitive to trypsinization of the cells and required an unoccupied high affinity lysine-binding site in the ligand. When plasminogen was added to the GM1380 cells, a line with high intrinsic plasminogen activator activity, the bound ligand was comprised of both plasminogen and plasmin. Urokinase, in catalytically active or inactive form, enhanced plasminogen binding to the two cell lines by 1.4-3.3-fold. Plasmin was the predominant form of the bound ligand when active urokinase was added, and preformed plasmin can also bind directly to the cells. Plasmin on the cell surface was also protected from its primary inhibitor, alpha 2-antiplasmin. These results indicate that the two cell lines possess specific binding sites for plasminogen and urokinase, and a family of widely distributed cellular receptors for these components may be considered. Endogenous or exogenous plasminogen activators can generate plasmin on cell surfaces, and such activation may provide a mechanism for arming cell surfaces with the broad proteolytic activity of this enzyme.     

Glutathione restores collagen degradation in TGF-beta-treated fibroblasts by blocking plasminogen activator inhibitor-1 expression and activating plasminogen

Transforming growth factor (TGF)-beta plays an important role in tissue fibrogenesis. We previously demonstrated that reduced glutathione (GSH) supplementation blocked collagen accumulation induced by TGF-beta in NIH-3T3 cells. In the present study, we show that supplementation of GSH restores the collagen degradation rate in TGF-beta-treated NIH-3T3 cells. Restoration of collagen degradation by GSH is associated with a reduction of type I plasminogen activator inhibitor (PAI)-1 expression/activity as well as recovery of the activities of cell/extracellular matrix-associated tissue-type plasminogen activator and plasmin. Furthermore, we find that NIH-3T3 cells constitutively express plasminogen mRNA and possess plasmin activity. Blockade of cell surface binding of plasminogen/plasminogen activation with tranexamic acid (TXA) or inhibition of plasmin activity with aprotinin significantly reduces the basal level of collagen degradation both in the presence or absence of exogenous plasminogen. Most importantly, addition of TXA or active PAI-1 almost completely eliminates the restorative effects of GSH on collagen degradation in TGF-beta treated cells. Together, our results suggest that the major mechanism by which GSH restores collagen degradation in TGF-beta-treated cells is through blocking PAI-1 expression, leading to increased PA/plasmin activity and consequent proteolytic degradation of collagens. This study provides mechanistic evidence for GSH's putative therapeutic effect in the treatment of fibrotic disorders.     

Plasminogen is a complement inhibitor

Plasminogen is a 92-kDa single chain glycoprotein that circulates in plasma as a zymogen and when converted to proteolytically active plasmin dissolves preformed fibrin clots and extracellular matrix components. Here, we characterize the role of plasmin(ogen) in the complement cascade. Plasminogen binds the central complement protein C3, the C3 cleavage products C3b and C3d, and C5. Plasminogen binds to C3, C3b, C3d, and C5 via lysine residues, and the interaction is ionic strength-dependent. Plasminogen and Factor H bind C3b; however, the two proteins bind to different sites and do not compete for binding. Plasminogen affects complement action in multiple ways. Plasminogen enhanced Factor I-mediated C3b degradation in the presence of the cofactor Factor H. Plasminogen when activated to plasmin inhibited complement as demonstrated by hemolytic assays using either rabbit or sheep erythrocytes. Similarly, plasmin either in the fluid phase or attached to surfaces inhibited complement that was activated via the alternative and classical pathways and cleaved C3b to fragments of 68, 40, 30, and 17 kDa. The C3b fragments generated by plasmin differ in size from those generated by the complement protease Factor I, suggesting that plasmin-mediated C3b cleavage fragments lack effector function. Plasmin also cleaved C5 to products of 65, 50, 30, and 25 kDa. Thus, plasmin(ogen) regulates both complement and coagulation, the two central cascade systems of a vertebrate organism. This complement-inhibitory activity of plasmin provides a new explanation why pathogenic microbes utilize plasmin(ogen) for immune evasion and tissue penetration.     

Cytosolic proteins contribute to surface plasminogen recruitment of Neisseria meningitidis

Plasminogen recruitment is a common strategy of pathogenic bacteria and results in a broad-spectrum surface-associated protease activity. Neisseria meningitidis has previously been shown to bind plasminogen. In this study, we show by several complementary approaches that endolase, DnaK, and peroxiredoxin, which are usually intracellular proteins, can also be located in the outer membrane and act as plasminogen receptors. Internal binding motifs, rather than C-terminal lysine residues, are responsible for plasminogen binding of the N. meningitidis receptors. Recombinant receptor proteins inhibit plasminogen association with N. meningitidis in a concentration-dependent manner. Besides binding purified plasminogen, N. meningitidis can also acquire plasminogen from human serum. Activation of N. meningitidis-associated plasminogen by urokinase results in functional activity and allows the bacteria to degrade fibrinogen. Furthermore, plasmin bound to N. meningitidis is protected against inactivation by alpha(2)-antiplasmin.     

Tissue plasminogen activator and urokinase mediate the binding of Glu-plasminogen to plasma fibrin I. Evidence for new binding sites in plasmin-degraded fibrin I

The effect of tissue plasminogen activator (TPA) or urokinase on the specific binding of human Glu-plasminogen to fibrin I formed in plasma by clotting with Reptilase was studied using 125I-plasminogen and 131I-fibrinogen. In the absence of TPA, small amounts of plasminogen were bound to fibrin I. TPA induced binding of plasminogen to plasma fibrin I that was dependent upon the concentrations of TPA and plasminogen as well as upon the time of incubation. Plasminogen binding occurred in association with fibrin clot lysis and the formation in the clot supernatant of alpha 2-plasmin inhibitor-plasmin complexes. Urokinase also induced binding of plasminogen to plasma fibrin I that was concentration- and time-dependent. The molecular form of plasminogen bound to the fibrin I plasma clot was identified as Glu-plasminogen by dodecyl sulfate-polyacrylamide gel electrophoresis and by fast performance liquid chromatography. Further studies demonstrated that fibrin I formed from fibrinogen that had been progressively degraded by plasmin-bound Glu-plasminogen. The mole ratio of plasminogen bound increased with the time of plasmin digestion. Glu-plasminogen did not bind to fibrin I formed from fibrinogen progressively digested by human leukocyte elastase, thereby demonstrating the specificity of plasmin. These studies demonstrate that plasminogen activators regulate the binding of Glu-plasminogen to fibrin I by catalyzing plasmin-mediated modifications in the fibrin substrate.     

Role of cell-surface lysines in plasminogen binding to cells: identification of alpha-enolase as a candidate plasminogen receptor.

Plasminogen binding to cell surfaces results in enhanced plasminogen activation, localization of the proteolytic activity of plasmin on cell surfaces, and protection of plasmin from alpha 2-antiplasmin. We sought to characterize candidate plasminogen binding sites on nucleated cells, using the U937 monocytoid cell as a model, specifically focusing on the role of cell-surface proteins with appropriately placed lysine residues as candidate plasminogen receptors. Lysine derivatives with free alpha-carboxyl groups and peptides with carboxy-terminal lysyl residues were effective inhibitors of plasminogen binding to the cells. One of the peptides, representing the carboxy-terminal 19 amino acids of alpha 2-antiplasmin, was approximately 5-fold more effective than others with carboxy-terminal lysines. Thus, in addition to a carboxy-terminal lysyl residue, other structural features of the cell-surface proteins may influence their affinity for plasminogen. Affinity chromatography has been used to isolate candidate plasminogen receptors from U937 cells. A major protein of Mr 54,000 was recovered and identified as alpha-enolase by immunochemical and functional criteria. alpha-Enolase was present on the cell surface and was capable of binding plasminogen in ligand blotting analyses. Plasminogen binding activity of a molecular weight similar to alpha-enolase also was present in a variety of other cell types. Carboxypeptidase B treatment of alpha-enolase abolished its ability to bind plasminogen, consistent with the presence of a C-terminal lysyl residue. Thus, cell-surface proteins with carboxy-terminal lysyl residues appear to function as plasminogen binding sites, and alpha-enolase has been identified as a prominent representative of this class of receptors.     

Plasminogen detection in oocytes and plasminogen activator activities in the porcine oviduct during the estrous cycle

Plasminogen activators (PAs) are highly specific serine proteases that convert the extracellular zymogen plasminogen into the active proteinase plasmin. Plasminogen-dependent proteolytic activity was detected by zymography both in the tissue membrane fraction of oviducts and in the oviductal flushing obtained at the preovulatory (Pre-Ov), postovulatory (Post-Ov) and mid-luteal (Mid-L) stages of the estrous cycle. A main proteolytic band, with a relative mobility similar to a human melanoma cell tissue-type plasminogen activator (t-PA), was found in all samples. Two additional components were observed in Pre-Ov and Post-Ov oviductal flushing but not in the tissue membrane fraction. In the oviductal flushing the PA activity was significantly higher in the Post-Ov stage than in the Pre-Ov one. Both urokinase-type plasminogen activator (u-PA, 50 kDa) and t-PA (72 kDa) were detected by Western blot; they showed differences in their relative concentration between Post-Ov and Pre-Ov oviductal flushing. The main PA substrate, plasminogen, was detected by indirect immunofluorescence in the cumulus cell extracellular matrix (ECM) and oocyte zona pellucida (ZP). In denuded oocytes, plasminogen was also detected on the surface of the plasma membrane. It is possible that oviductal PAs may act on the plasminogen present in the cumulus cell ECM and ZP; consequently, the generated plasmin could be involved in the rebuilding or degradation of these oocyte structures during fertilization or early development.     

Analysis of plasminogen-binding M proteins of Streptococcus pyogenes

Group A streptococci are common human pathogens that cause a variety of infections. They express M proteins which are important cell wall-bound type-specific virulence factors. We have found that a set of strains, associated primarily with skin infections, express M proteins that bind plasminogen and plasmin with high affinity. The binding is mediated by a 13-amino-acid internal repeated sequence located in the N-terminal surface-exposed portion of these M proteins. This sequence binds to kringle 2 in plasminogen, a domain that is not involved in the interaction with streptokinase, a potent group A streptococcal activator of plasminogen. It could be demonstrated that plasminogen, absorbed from plasma by growing group A streptococci expressing the plasminogen-binding M proteins, could be activated by exogenous and endogenous streptokinase, thereby providing the bacteria with a surface-associated enzyme that could act on the tissue barriers in the infected host.