Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:3.4.21.37 (neutrophil elastase)
 4,078 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Limited proteolysis of T-kininogen by heterologous and homologous endopeptidases (bovine trypsin, human leukocyte elastase, rat submaxillary gland endopeptidase k, and rat mast cell chymase) produced similar fragmentation. Amino-terminal sequence analysis of whole T-kininogen lysates and purified proteolytic fragments identified four susceptible regions which contained all the preferential cleavage sites for these proteinases. Two of these susceptible regions were close to the junction between heavy chain cystatin-like domains, the third was in the kinin-containing region, and the fourth was close to the carboxyl terminus of the T-kininogen light chain. There was only one primary site for each proteinase in the kinin-containing region, which explains why catalytic amounts of these proteinases did not release immunoreactive kinin from this kininogen. However, preferential cleavage of T-kininogen close to the junction between cystatin-like domains released fragments which, provided they included cystatin-like domains 2 and/or 3, strongly inhibited papain and cathepsin L. The fragments were inhibitory even when parts of the amino-terminal ends of the domains were lacking. The highly conserved glycyl residue, thought to be involved in the inhibitory reactive site of cystatin-like inhibitors, was not required in purified domain 3 for inhibition of cathepsin L.
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PMID:Limited proteolysis of T-kininogen (thiostatin). Release of comparable fragments by different endopeptidases. 264 33

Plasma kallikrein has been shown to aggregate human neutrophils and release human neutrophil elastase. However, neutrophils resuspended in factor XII-deficient plasma released only 30% of the elastase compared with normal plasma. Isolated human neutrophils were aggregated in a concentration-dependent fashion by 0.06 to 0.6 U/mL factor XIIa (0.022 to 0.22 mumol/L). Factor XIIa (0.1 to 1.0 U/mL) also induced neutrophil degranulation as evidenced by a concentration-dependent release of the specific granule protein, lactoferrin, and azurophilic granule protease, elastase. The release of neutrophil elastase was biphasic, reaching 40% of maximum at 15 seconds with maximal release by 90 minutes. The active site of factor XIIa was required, since the synthetic inhibitor, D-Pro-Phe-Arg-CH2Cl, which reacts with an essential histidine, and the natural plasma inhibitor, Cl-inhibitor, which interacts with the critical serine, both inhibit by more than 90% the release of elastase. The heavy chain is also required, since factor XII fragments failed to aggregate neutrophils or stimulate degranulation. Factor XIIa (0.6 U/mL) can completely correct the defect in elastase release evident in factor XII-deficient plasma. These studies demonstrate that factor XIIa, at concentrations potentially obtainable in plasma in disease states, can activate neutrophils, and thus may participate in the inflammatory response.
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PMID:Purified plasma factor XIIa aggregates human neutrophils and causes degranulation. 348 86

Mouse macrophage elastase, a metalloproteinase secreted by inflammatory macrophages, catalyzed the limited proteolysis of selected subclasses of mouse immunoglobulins, including monomeric IgG2a, IgG3, and some forms of IgG2b. Mouse IgG1 was resistant to elastase degradation; however, human IgG1 was degraded. IgG3 in immune complexes was cleaved in a manner similar to that of monomeric IgG3. Degradation by macrophage elastase was limited to the heavy chain, resulting in products that did not compete for binding to the macrophage Fc receptor. Macrophage elastase usually produced a pepsin-like rather than a papain-like pattern of proteolysis, resulting in the release of F(ab')2 and Fc' subfragments. This degradation of IgG differed from the papain-like cleavage of IgG by granulocyte elastase. Macrophage elastase degraded papain-generated Fc fragments of IgG2a into multiple fragments. Therefore, macrophage elastase at concentrations found in culture medium has the potential to regulate some aspects of cellular events associated with immunoglobulins.
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PMID:Selective proteolysis of immunoglobulins by mouse macrophage elastase. 622 Jan 7

Recently it has been demonstrated that human antibody fragments with binding activities against self antigens can be isolated from repertoires of rearranged V genes from non-immunized humans. We have applied phage display technology to study the B cell repertoire for antibody activity against neutrophil cytoplasmic antigens. These antibodies may play an important role in Wegener's granulomatosis (WG) and related forms of vasculitides. Autoantibodies in patients with WG are directed against proteinase 3. The immunodominant antigen in other forms of vasculitis is myeloperoxidase, but the B cell response can also be directed against other neutrophil enzymes, e.g. lysozyme, human neutrophil elastase, lactoferrin and cathepsin G. We show here that anti-self reactivity against neutrophil cytoplasmic antigens can be detected in the rearranged V gene repertoire of healthy individuals and that the reactivity can be directed against structural related epitopes which are present on different neutrophil cytoplasmic antigens. The scFv with binding activities were sequenced and the V gene usage, the level of somatic mutations and the immunoserological characteristics of the antibody fragments are discussed. Further evidence is presented that antibody fragments consisting only of a heavy chain variable domain can recognize neutrophil cytoplasmic antigens in a specific manner. These single-domain antibody fragments were used in experiments designed to establish the relative role of the light chain variable domains in antigen binding.
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PMID:Molecular characteristics of anti-self antibody fragments against neutrophil cytoplasmic antigens from human V gene phage display libraries. 853 74

The effect of human neutrophil elastase (HNE) on human factor V (F.V) or alpha-thrombin-activated human factor V (F.Va) was studied in vitro by prothrombinase assays, sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE), and NH2-terminal sequence analysis. Incubation of F.V (600 nmol/L) with HNE (2 nmol/L) in the presence of Ca2+ resulted in a time-dependent increase in its cofactor activity. In contrast, treatment of F.Va (600 nmol/L) with HNE (60 nmol/L) in the presence of Ca2+ resulted only in a time-dependent decrease in its cofactor activity. Under the conditions of these experiments, the maximum extent of F.V activation accomplished by incubation with HNE was approximately 65% to 70% of that observed with alpha-thrombin in presence of Ca2+. The extent of both the HNE-dependent enhancement in F.V cofactor activity and the HNE-dependent decrease in F.Va cofactor activity was not influenced by the addition of phosphatidylcholine/phosphatidylserine (PCPS) vesicles (50 micromol/L). The HNE-derived cleavage products of F.V, which correlated with increased cofactor activity, as demonstrated by SDS-PAGE under reducing conditions, were different from those generated using alpha-thrombin. Treatment of F.V (600 nmol/L) with HNE (2 nmol/L) in the presence of Ca2+ resulted in the production of three closely spaced doublets of: 99/97, 89/87, and 76/74 kD whose appearance over time correlated well with the increased cofactor activity as judged by densitometry. Treatment of F.Va (600 nmol/L) with HNE (60 nmol/L) in the presence of Ca2+ resulted in the cleavage of both the 96 kD heavy chain and the 74/72 kD light chain into products of: 56, 53, 35, 28, 22, and 12 kD. Although densitometry indicated that both the heavy and light chains of F.Va were hydrolyzed by HNE, cleavage of the 96 kD heavy chain was more extensive during the time period (10 to 30 minutes) of the greatest loss of F.Va cofactor activity. NH2-terminal sequence analysis of F.V treated with HNE indicated cleavage at Ile819 and Ile1484 under conditions during which the procofactor expressed enhanced cofactor activity in the prothrombinase complex. NH2-terminal sequence analysis of F.Va treated with HNE indicated cleavage at Ala341, Ile508, and Thr1767 under conditions, which the cofactor became inactivated, as measured by prothrombinase activity. The activation and inactivation cleavage sites are close to those cleaved by the physiological activator and inactivator of F.V and F.Va, namely alpha-thrombin (Arg709 and Arg1545) and Activated Protein C (APC) (Arg306 and Arg506), respectively. These results indicate that HNE can generate proteolytic products of F.V, which initially express significantly enhanced procoagulant cofactor activity similar to that observed following activation with alpha-thrombin. In contrast, HNE treatment of F.Va resulted only in the loss of its cofactor activity, but again, this is similar to that observed following inactivation by APC.
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PMID:Human neutrophil elastase activates human factor V but inactivates thrombin-activated human factor V. 924 37

We have previously demonstrated a low-affinity (0.8 microM, non-covalent complex formation between high-molecular-mass kininogen (HK) and plasminogen (Plg) which prevented Plg interaction with glioma and endothelial cells. We have now extended our previous observations by exploring the potential complex formation between Plg and low-molecular-mass kininogen (LK) and between LK and HK with Plg cleaved with human neutrophil elastase (HNE). Plg cleavage by HNE (PlgHNE) yielded kringles 1-3, kringle 4 and mini-plasminogen. PlgHNE was subjected to SDS/PAGE under non-reducing conditions, followed by western blotting, and incubated with either 125I-HK or 125I-LK. Autoradiograms revealed that 125I-HK bound to miniplasminogen and to kringles 1-3 but not to kringle 4 and the presence of 10 mM 6-aminohexanoic acid (Ahx) disrupted only the interaction with kringles 1-3. In contrast, 125I-LK bound to miniplasminogen but not to kringles 1-3 or 4 and Ahx had no effect at all. The complex formation of either HK (0.67 microM) or LK (3 microM) with Plg (1.5 microM) did not affect its conversion to plasmin by tissue plasminogen activator (t-PA) (10 U/ml) in the presence of a tissue plasminogen stimulator (0.14 microM). However, the rate of conversion of plasminogen to plasmin by t-PA was affected when platelets were added to the reaction mixture. Since HK (0.83 microM) has been shown to inhibit plasmin-induced platelet aggregation, we investigated whether this inhibitory property is found within the heavy chain shared by HK and LK. We found that LK inhibited plasmin-induced platelet aggregation, but a 4-fold molar excess was required when compared to HK. Compared to plasmin, 3-5-fold molar excess of miniplasmin is required to induce platelet aggregation, indicating the important role of kringles 1-3 for plasmin interactions with these cells. These results indicate that HK and LK-mediated inhibition of plasmin-induced platelet aggregation is likely due to complex formation with kringle 5 without interfering with plasmin's active site. We found an additional interaction between HK and kringles 1-3 enhancing the inhibitory effect, presumably by interfering with plasmin's interaction with platelets. This HK and LK-associated modulation of plasmin-induced platelet aggregation may serve as a template to develop synthetic peptides as novel therapeutic agents to prevent some of the plasmin-associated thrombocytopenia seen during thrombolytic therapy.
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PMID:High-molecular-mass and low-molecular-mass kininogens block plasmin-induced platelet aggregation by forming a complex with kringle 5 of plasminogen/plasmin. 942 7

Factor V (FV) is a large (2,196 amino acids) nonenzymatic cofactor in the coagulation cascade with a domain organization (A1-A2-B-A3-C1-C2) similar to the one of factor VIII (FVIII). FV is activated to factor Va (FVa) by thrombin, which cleaves away the B domain leaving a heterodimeric structure composed of a heavy chain (A1-A2) and a light chain (A3-C1-C2). Activated protein C (APC), together with its cofactor protein S (PS), inhibits the coagulation cascade via limited proteolysis of FVa and FVIIIa (APC cleaves FVa at residues R306, R506, and R679). The A domains of FV and FVIII share important sequence identity with the plasma copper-binding protein ceruloplasmin (CP). The X-ray structure of CP and theoretical models for FVIII have been recently reported. This information allowed us to build a theoretical model (994 residues) for the A domains of human FV/FVa (residues 1-656 and 1546-1883). Structural analysis of the FV model indicates that: (a) the three A domains are arranged in a triangular fashion as in the case of CP and the organization of these domains should remain essentially the same before and after activation; (b) a Type II copper ion is located at the A1-A3 interface; (c) residues R306 and R506 (cleavage sites for APC) are both solvent exposed; (d) residues 1667-1765 within the A3 domain, expected to interact with the membrane, are essentially buried; (e) APC does not bind to FVa residues 1865-1874. Several other features of factor V/Va, like the R506Q and A221V mutations; factor Xa (FXa) and human neutrophil elastase (HNE) cleavages; protein S, prothrombin and FXa binding, are also investigated.
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PMID:Structural investigation of the A domains of human blood coagulation factor V by molecular modeling. 965 35

A comparative in vitro survey of physiologically relevant human and microbial proteinases defined a number of enzymes that induced specific hinge domain cleavage in human IgG1. Several of these proteinases have been associated with tumor growth, inflammation, and infection. A majority of the identified proteinases converted IgG to F(ab')(2), and a consistent feature of their action was a transient accumulation of a single-cleaved intermediate (scIgG). The scIgG resulted from the relatively rapid cleavage of the first hinge domain heavy chain, followed by a slower cleavage of the second chain to separate the Fc domain from F(ab')(2). Major sites of enzymatic cleavage were identified or confirmed from the mass of the F(ab')(2) or Fab fragments and/or the amino-terminal amino acid sequence of the Fc for each enzyme including human matrix metalloproteinases (MMPs) 3 and 12, human cathepsin G, human neutrophil elastase (Fab), staphylococcal glutamyl endopeptidase I and streptococcal immunoglobulin-degrading enzyme (IdeS). The cleavage sites in IgG1 by MMP-3, cathepsin G and IdeS were used to guide the synthesis of peptide analogs containing the corresponding carboxy-termini to be used as immunogens in rabbits. Rabbit antibodies were successfully generated that showed selective binding to different human F(ab')(2)s and other hinge-cleavage fragments, but not to intact IgG. In Western blotting studies of synovial fluids from individuals with rheumatoid arthritis, the rabbit antibodies yielded patterns consistent with the presence of endogenous IgG fragments including F(ab')(2) and the single-cleaved IgG intermediate. The detection in synovial fluid of IgG fragments similar to those observed in the in vitro biochemical studies suggests that proteolysis of IgG may contribute to localized immune dysfunction in inflammatory environments.
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PMID:Proteolysis of purified IgGs by human and bacterial enzymes in vitro and the detection of specific proteolytic fragments of endogenous IgG in rheumatoid synovial fluid. 1815 32

Mechanisms of inflammation and coagulation are linked through various pathways. Human neutrophil elastase (HNE), can bind to activated platelets, might be localised on platelet membranes that provide negatively-charged phospholipid essential for the optimum function of tenase complex. In this study, we examined the effect of HNE on factor (F)VIII. FVIII activity was rapidly diminished in the presence of HNE and was undetectable within 10 minutes. The inactivation rate was ~8-fold greater than that of activated protein C (APC). This time-dependent inactivation was moderately affected by von Willebrand factor. HNE proteolysed the heavy chain (HCh) of FVIII into two terminal products, A11-358 and A2375-708, by limited proteolysis at Val358, Val374, and Val708. Cleavage at Val708 was much slower than that at Val358 in the >90-kDa A1-A2-B compared to the 90-kDa A1-A2. The 80-kDa light chain (LCh) was proteolysed to 75-kDa product by cleavage at Val1670. HNE-catalysed FVIIIa inactivation was markedly slower than that of native FVIII (by ~25-fold), due to delayed cleavage at Val708 in FVIIIa. The inactivation rate mediated by HNE was ~8-fold lower than that by APC. Cleavages at Val358 and Val708 were regulated by the presence of LCh and HCh, respectively. In conclusion, HNE-catalysed FVIII inactivation was associated with the limited-proteolysis that led to A11-358, A2375-708, and A3-C1-C21671-2332, and subsequently to critical cleavage at Val708. HNE-related FVIII(a) reaction might play a role in inactivation of HNE-induced coagulation process, and appeared to depend on the amounts of inactivated FVIII and active FVIIIa which is predominantly resistant to HNE inactivation.
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PMID:Mechanisms of human neutrophil elastase-catalysed inactivation of factor VIII(a). 2147 77