Purification of plasminogen from human granulocytes using DEAE-Sephadex column chromatography

Prokopowicz, J

doi:10.1016/0005-2795(68)90262-6

Cited by 20 publications

(12 citation statements)

References 11 publications

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“…This observation is consistent with the results of other studies (11)(12)(13) in which plasmin activity and the antigenic determinants of plasmin were not demonstrable in leukocytes. In contrast, Prokopowicz not only found evidence for the presence of the plasminogen system in leukocytes but also isolated plasminogen from leukocytes (14,17). Our observations do not resolve these apparent inconsistencies; but in preliminary observations, utilizing sensitive radioimmunochemical approaches (25,29), we have detected very low levels of plasmin-related antigens within several leukocyte preparations (nanogram levels per 10' cells).…”

Section: Nonlytic Release Of Fibrinolytic Proteases From Leukocytescontrasting

confidence: 77%

“…The gels were subjected to electrophoresis at 8 mA/gel and stained with Coomassie brilliant blue. Molecular weights of constituent chains of the various digests were estimated in the reduced system utilizing serum albumin (68,000), the heavy (50,000) and light chains (23,500) of IgG, ovalbumin (43,000), pepsin (35,000), trypsin (23,300), and myoglobulin (17,200) as standards.…”

Section: Methodsmentioning

confidence: 99%

“…plasminogen and its proactivator, may be constituents of leukocytes (14,17), a number of studies have demonstrated within the cells a fibrinolytic system distinct from plasmin. This conclusion is based on the differential sensitivity of the systems to inhibitors, as well as comparisons of the molecular characteristics of the enzymes (11)(12)(13)(14).…”

Section: Introductionmentioning

confidence: 99%

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An alternative pathway for fibrinolysis. I. The cleavage of fibrinogen by leukocyte proteases at physiologic pH.

Plow¹,

Edgington²

1975

J. Clin. Invest.

209

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A B S T R A C T An alternative fibrinolytic system, active at physiological pH, is present in peripheral blood leukocytes. The fibrinolytic proteases localized predominantly in the leukocyte granules are capable of degrading both fibrinogen and fibrin, and plasmin activity does not contribute significantly to this proteolytic event.The specificity of the alternative fibrinolytic proteases for fibrinogen and the characteristics of the derivative cleavage fragments are clearly distinguishable from the classical plasmin system. The high molecular weight derivatives of fibrinogen, generated by the alternative system, under physiological conditions, are larger than the plasmin-generated X fragment, exhibit immunoelectrophoretic mobility comparable to native fibrinogen, and are not coagulable by thrombin. Analysis of the constituent polypeptide chains of the fragments reveals cleavage of the Aa, BP, and y chains of fibrinogen. The lower molecular weight derivatives of fibrinogen, generated by the alternative system, are structurally distinct from previously described fibrinogen degradation products and exhibit potent anticoagulant activity. This anticoagulant activity can be attributed to interference with normal fibrin polymerization. The proteases of the alternative fibrinolytic systems are actively secreted by leukocytes when stimulated to undergo a nonlytic release reaction. These results provide direct evidence for a fibrinolytic system resident in leukocyte granules that is associated with the leukocyte release reaction and is capable of generating unique fibrinogen cleavage fragments.

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Section: Nonlytic Release Of Fibrinolytic Proteases From Leukocytescontrasting

confidence: 77%

Section: Methodsmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

See 1 more Smart Citation

An alternative pathway for fibrinolysis. I. The cleavage of fibrinogen by leukocyte proteases at physiologic pH.

Plow¹,

Edgington²

1975

J. Clin. Invest.

209

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“…Further studies indicated that both fibrinogen and fibrin are susceptible to the action of leukocyte enzymes (3)(4)(5) and that the intrinsic enzymatic system of the leukocytes is distinct from plasmin (6)(7)(8)(9)(10). Apparently, intracellular plasminogen does not account for fibrinolysis, as only very low plasminogen levels have been detected in polymorphonuclear leukocytes (8,11,12).…”

Section: Introductionmentioning

confidence: 99%

Degradation products of fibrinogen by elastase-like neutral protease from human granulocytes. Characterization and effects on blood coagulation in vitro.

Gramse¹,

Bingenheimer²,

Schmidt³

et al. 1978

J. Clin. Invest.

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A B S T R A C T We investigated the effect of elastaselike neutral protease isolated from human granulocytes on human fibrinogen. Dependent on enzyme concentration and time of incubation, the elastase-like protease induced a progressive degradation of fibrinogen. Analysis of the remaining polypeptide chains showed a high susceptibility of the Aa-and low susceptibility of the y-chain of fibrinogen towards the proteolytic action of the enzyme. The split products were characterized by polyacrylamide gel electrophoresis and two-dimensional immunoelectrophoresis. They showed antigenic determinants of fibrinogen and of plasmin-induced proteolysis products D and E. The cleavage fragments isolated by gel chromatography had distinct molecular weights. Coagulability of fibrinogen by thrombin was inhibited according to the concentration of the protease and the time of incubation. Split products of fibrinogen with higher molecular weight prolonged the coagulation time of native fibrinogen, whereas low molecular weight fragments were ineffective.

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“…An elastase was purified and partially characterized by Janoff and Scherer [52], Janoff [53,55], J anoff and Basch [54], O hlsson and O lsson [56], Schmidt and H avermann [57], and recently by T ay lor and Crawford [58]. A chondromucoprotein-degrading activity was demonstrated in lysates of human PMN leukocyte granules [59], attribut able probably to the elastase [60], Fibrinolytic activity has been described in neutrophils, both in form of an active enzyme and a zymogen [61,62]. The latter was claimed to be identical to plasminogen of plasma [62].…”

Section: Discussionmentioning

confidence: 99%

Neutral Proteases of Human PMN Leukocytes with Kininogenase Activity

Movat¹,

Habal²,

Macmorine³

1976

Int Arch Allergy Immunol

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A neutral protease with kininogenase activity was isolated from human polymorphonuclear (PMN) leukocytes by cation exchange chromatography and gel filtration. The protease appears heterogeneous by cation exchange chromatography, isoelectric focusing and cationic disc gel electrophoresis, but homogeneous by gel filtration, sucrose density gradient ultracentrifugation, SDS-disc gel electrophoresis and immunoelectrophoresis. By carrying out the electrophoresis of the protease in acrylamide gels of varying concentrations, it was shown that they represent charge isomers. The protease was stable at pH 4–10, but labile to heat, being almost completely inactivated when incubated for 30 min at 70°C. It exhibited proteolytic activity between pH 5 and 9, being maximal at 7.5–8.5. The molecular weight of the PMN protease was estimated to be about 20,000 daltons by gel filtration in aqueous buffer and about 26,000–28,000 daltons by SDS-disc gel electrophoresis and gel filtration in Sepharose 6B in the presence of the dissociating agent guanidine HC1. Its sedimentation coefficient was about 2.7S. Corresponding to the charge heterogeneity, by isoelectric focusing, the kinin-generating and esterolytic activities of the PMN granule lysate focused between pH 6.0 and 11.5, whereas the isolated PMN protease focused between 10.0 and 11.8. With respect to kinin generation, caseinolysis, and alanine esterase activity, the protease was inhibited by DFP and certain chloromethyl ketone inhibitors, as well as the plasma protease inhibitory α₁-antitrypsin, α₂-macroglobulin and antithrombin III. Both bradykinin and a methionyl-lysyl-bradykinin-like peptide were generated from highly purified kininogens by a lysosomal lysate containing the PMN protease. However, this assay was done with a crude enzyme preparation which contains an aminopeptidase capable of converting lysyl-bradykinin or methionyl-lysyl-bradykinin to bradykinin. When injected intradermally, the protease induced hyperemia, hemorrhage, and moderate enhancement of vascular permeability, but the mixture of the protease and kininogen induced a marked enhancement of vascular permeability.

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Purification of plasminogen from human granulocytes using DEAE-Sephadex column chromatography

Cited by 20 publications

References 11 publications

An alternative pathway for fibrinolysis. I. The cleavage of fibrinogen by leukocyte proteases at physiologic pH.

An alternative pathway for fibrinolysis. I. The cleavage of fibrinogen by leukocyte proteases at physiologic pH.

Degradation products of fibrinogen by elastase-like neutral protease from human granulocytes. Characterization and effects on blood coagulation in vitro.

Neutral Proteases of Human PMN Leukocytes with Kininogenase Activity

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