H Hörmann

Max Planck Institute of Biochemistry, München, Bavaria, Germany

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Publications (38)56.6 Total impact

  • H Hörmann, H Richter, V Jelinić
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    ABSTRACT: Previous experiments had shown that the free N-terminal fibronectin 30-kDa-domain mediates binding of soluble 125I-fibrin to transamidase-coated polystyrene beads (Hörmann et al., Biol. Chem. Hoppe-Seyler 368, 669-674, 1987). Now, the formation of covalent adducts of the N-terminal fragment with fibrin peptide chains is demonstrated. Binding of soluble 125I-fibrin was performed in presence of N-terminal fibronectin 30-kDa or 70-kDa fragments. The material adsorbed was removed from the beads under reducing conditions and analysed by dodecylsulfate gel electrophoresis followed by autoradiography. The 30-kDa fragment gave rise to bands of 80 kDa and 180-200 kDa which were lacking in the products of the 70-kDa compound. Instead, they showed bands at 120 kDa and ca. 280 kDa. Evidently, those bands represented covalent adducts of fibrin peptide chains or their dimers with the 30-kDa or the 70-kDa fragment, respectively. In addition, dimeric gamma-chains and alpha-chain polymers of fibrin were present indicating partial polymerization of bead-attached fibrin.
    Biological chemistry Hoppe-Seyler 07/1991; 372(6):427-30. · 2.68 Impact Factor
  • H Hörmann, V Jelinić, H Richter
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    ABSTRACT: Centrifuged human platelets bound soluble 125I-labelled fibrin, mediated by a plasma factor. Binding was inhibited by D-phenylalanyl-L-prolyl-L-arginyl- chloromethane (PPACK), which specifically blocks thrombin. As the binding-promoting principle was adsorbed to barium citrate, it was tentatively characterized as prothrombin, suggesting that it might be converted to thrombin at the cell surface. The peptide GRGDSP failed to inhibit binding, thus eliminating the glycoprotein IIb/IIIa complex as a receptor. Most likely, a thrombin - fibrin complex is recognized by a cell receptor, possibly protease-nexin I. In a platelet concentrate, the cells also internalized 125I-labelled fibrin, providing evidence that platelets are involved in the clearance of circulating fibrin - monomer complexes. Engulfment was again inhibited by PPACK or hirudin but not by an antibody against the glycoprotein IIb/IIIa complex.
    Blood Coagulation and Fibrinolysis 11/1990; 1(4-5):547-9. · 1.38 Impact Factor
  • Helmut Hörmann, Hartmut Richter, V Jelinić
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    ABSTRACT: Binding of soluble 125I-fibrin to platelets was investigated with thrombocytes separated by gelfiltration or by sedimentation as well as in a thrombocyte concentrate. Gelfiltered platelets failed to retain 125I-fibrin within 16 hours unless they had been pretreated with factor XIIIa. In addition, a 30 kDa-component derived from the N-terminal fibronectin domain was required as a mediator. Platelets isolated by sedimentation bound some 125I-fibrin even in the absence of those cofactors. The 30 kDa-component improved binding and only this increase was sensitive to putrescine inhibition. Evidently, centrifuged platelets unlike gelfiltered ones express two pathways of fibrin binding. In a thrombocyte concentrate with platelets in their plasmatic environment 125I-fibrin was partially internalized. Engulfed radioactivity was detectable only for a limited period between 4-6 hours after substrate application suggesting that 125I-fibrin was intracellularly degraded followed by release of the fragments. The 30 kDa-component promoted internalization, while factor XIIIa improved the capacity. Thrombin inhibitors suppressed the uptake.
    Thrombosis Research 05/1990; 58(2):175-83. · 2.43 Impact Factor
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    ABSTRACT: The plasma free N-terminal fibronectin 30-kDa domain was measured in 44 type 1 diabetic patients and in 20 healthy subjects. A significantly raised mean concentration of a free N-terminal fibronectin 30-kDa domain was found in plasma of diabetic patients with proliferative retinopathy as compared with healthy persons (P less than 0.001). A positive correlation was observed between free N-terminal fibronectin 30-kDa domain and von Willebrand factor in plasma of all examined subjects (r = 0.62, P less than 0.01). A similar correlation was present between 30-kDa domain and albuminuria (r = 0.56, P less than 0.01). However, no relationship was found between fibronectin 30-kDa domain and control of diabetes as assessed by fructosamine concentration. The free N-terminal fibronectin 30-kDa domain may be used as a marker of actual endothelial cell dysfunction in diabetes.
    European Journal of Clinical Investigation 05/1990; 20(2):171-6. · 2.83 Impact Factor
  • H Hörmann, V Jelinić, H Richter
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    ABSTRACT: Previous experiments (Hörmann, H. & Jelinić, V. (1980) Hoppe-Seyler's Z. Physiol. Chem. 361, 379-387) had shown that heparin promoted the binding of plasma fibronectin to peritoneal macrophages of guinea pigs. The present data reveal that this effect only takes place at higher fibronectin concentrations indicating cooperative processes, most likely association of fibronectin at the cell surface. An unspecific precipitation of fibronectin by heparin was prevented by calcium in the medium. The accumulation at the cell surface was inhibited by the following fibronectin fragments: N-terminal 30 kDa and 70 kDa containing a potential self-association site and a transamidase-reactive site; central 95 kDa which comprised a negatively charged region possibly involved in self-association as well as the so-called alternative cell-binding site, but was lacking the cell-binding Arg-Gly-Asp sequence; heparin-binding 37-kDa and 60-kDa fragments. All these domains and sites, therefore, were potentially important in the assembly process at the cell surface. A peptide comprising the sequence Arg-Gly-Asp was ineffective pointing against an involvement of this fibronectin cell-binding site in the overall process. Macrophages of older animals were less capable of accumulating fibronectin under the reaction conditions. Their capability was improved after preincubation with activated plasma transglutaminase (coagulation factor XIIIa) suggesting that a cell-attached transamidase might be important for the assembly process.
    Biological chemistry Hoppe-Seyler 07/1989; 370(7):691-8. · 2.68 Impact Factor
  • J Skrha, Hartmut Richter, Helmut Hörmann
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    ABSTRACT: The free N-terminal 30-kDa domain of the fibronectin subunit chains had previously been shown to mediate binding of soluble fibrin to phagocytic cells. In order to demonstrate whether the fragment is available in plasma in a suitable concentration, an indirect immunoassay procedure for its quantitative evaluation was developed. The free form of the 30-kDa domain was separated from fibronectin and the bulk of the plasma proteins by two-step affinity chromatography on gelatin- and heparin-Sepharose. In the eluate of the heparin-Sepharose the 30-kDa fragment was determined by its capacity to inhibit the immune reaction between a specific antiserum and the 30-kDa fragment immobilized on microtiter wells. The procedure offered reproductibility comparable with other immunoassays (coefficient of variation 4.0 to 8.0%); the lowest amount of detectable 30-kDa fragment was 0.1 microgram/ml. In human plasma this method detected for the first time ca. 5 micrograms/ml 30-kDa fragment. This concentration is in the range required for binding of fibrin to cells.
    Analytical Biochemistry 10/1988; 173(2):228-34. · 2.31 Impact Factor
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    ABSTRACT: Gelfiltered unstimulated human platelets neither bound 125-I-fibrinogen nor 125-I-fibrin. Fibrin-binding was, however, stimulated by N-terminal fibronectin 30 kD-and 70 kD-fragments while fibronectin was ineffective. The 30 kD-fragment also stimulated some platelet preparations to bind fibrinogen which, however, was suppressed by minute amounts of the thrombin inhibitor PPACK. PPACK hardly influenced fibrin-binding. Fragment-promoted fibrinogen-binding was also inhibited by a monoclonal antibody recognizing the membrane glycoprotein IIb/IIIa complex known to act as fibrinogen receptor. This antibody failed to influence fragment-stimulated fibrin-binding giving evidence that fibrinogen and fibrin were retained by different receptors. In contrast to 125-I-fibrin its plasmin-derived and 125-I-labelled fragment X was not recognized by the platelets in presence of the fibronectin 30 kD-fragment. Fragment-stimulated binding of 125-I-fibrin showed a lag phase and was completely inhibited by 0.25 mM putrescine as well as by 1 mM EDTA or 0.1 mM N-ethylmaleinimide. Evidently, a cell-attached transamidase was involved in fibrin-binding possibly by forming a ternary complex with fibrin and the fibronectin fragment.
    Thrombosis Research 09/1988; 51(3):283-93. · 2.43 Impact Factor
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    ABSTRACT: Polystyrene beads coated with thrombin-activated Factor XIII (plasma transglutaminase, plasma transamidase) retained soluble 125I-fibrin, if a 30-kDa fragment of fibronectin was present. The fragment was obtained by proteolytic cleavage of plasma fibronectin with trypsin, but was also available with plasmin or thrombin. It represented a fibrin-binding domain at the N-terminus of each of the two subunit chains and contained close to its amino end a transamidase-reactive site. Fibronectin was unable to mediate the binding of 125I-fibrin to Factor XIIIa-coated beads. 125I-fibrinogen was hardly recognized by the beads even in presence of the fibronectin fragment. The relatively slow binding of 125I-fibrin was inhibited by 0.15 mM putrescine or by a pretreatment of the coated beads with EDTA or N-ethylmaleinimide indicating the involvement of a transamidation in the binding reaction. Immobilization of 125I-fibrin in presence of the fibronectin fragment is assumed to require a covalent cross-linking of the two ligands at the immobilized transamidase giving rise to a product which is retained strongly. The possibility is discussed that a surface-attached transamidase might act as a fibrin receptor which requires the fibronectin fragment as a cofactor.
    Biological chemistry Hoppe-Seyler 07/1987; 368(6):669-74. · 2.68 Impact Factor
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    ABSTRACT: Immunogold labeling was used to localize the core protein of small dermatan sulfate proteoglycan (DS-PG) on the surface of cultured human fibroblasts. At 4 degrees C, DS-PG core protein was uniformly distributed over the cell surface. At 37 degrees C, gold particles either became rearranged in form of clusters or remained associated with fibrils. Double-label immunocytochemistry indicated the co-distribution of DS-PG core protein and fibronectin in the fibrils. In an enzyme-linked immunosorbent assay, binding of DS-PG from fibroblast secretions and of its core protein to fibronectin occurred at pH 7.4 and at physiological ionic strength. Larger amounts of core protein than of intact proteoglycan could be bound. Fibronectin peptides containing either the heparin-binding domain near the COOH-terminal end or the heparin-binding NH2 terminus were the only fragments interacting with DS-PG and core protein. Competition and replacement experiments with heparin and dermatan sulfate suggested the existence of adjacent binding sites for heparin and DS-PG core protein. It is hypothesized that heparan sulfate proteoglycans and DS-PG may competitively interact with fibronectin.
    The Journal of Cell Biology 07/1987; 104(6):1683-91. · 9.69 Impact Factor
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    ABSTRACT: Proteolytic fragments from the N-terminus of the fibronectin subunit chains were shown to mediate the binding of 125-I-fibrin to macrophages. With increasing molecular weight of the fragments, binding activity decreased and intact plasma fibronectin was inactive. Fibrin binding to macrophages was a time dependent reaction and proceeded considerably faster than binding of fibrinogen. The binding reaction was inhibited by putrescine suggesting the involvement of a transamidase. Pericellular transamidase was demonstrated on macrophages by incorporation of 14-C-putrescine into fibronectin 30 kD-fragment. Expression of this enzyme appeared to be rate-limiting for the binding reaction which was accelerated after loading the cells with placental transamidase.
    Thrombosis Research 05/1987; 46(1-46):39-50. · 2.43 Impact Factor
  • H Hörmann, H Richter
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    ABSTRACT: Fibronectin, consisting of two threadlike subunits connected close to their C-terminal ends, exists in a soluble and a fibrous form. Models are presented that propose distinct foldings of the rodlike subunits in the soluble dimer and an extended arrangement in the aggregates. The proposed conformations are based on the analysis of electrostatic attractions and noncovalent affinities between domains. Electrofocusing of proteolytic fragments revealed a sequence of four domains with alternating charges in the N-terminal half of each subunit and two domains with opposite charges close to the C-terminus. Complementary sites with affinity to each other were localized by radioimmuno-binding assay in the gelatin-binding and in the subsequent DNA-binding domain of the N-terminal tetra-domain sequence. Claiming that in soluble fibronectin electrostatic attractions and noncovalent affinities should be neutralized within the molecule resulted in the construction of a conformation with backfolded subunits, each containing an extra loop in which domains with complementary affinity sites are saturated by each other. The model is in accord with hydrodynamic and electronmicroscopic data. There is, however, an alternative folding in which electrostatic and noncovalent affinity sites in the N-terminal half of each subunit are saturated by an interchain interaction within the molecule. Consequently, a rearrangement of the molecule without significant shape change cannot be excluded. In the aggregated form, the N-terminal tetra-domain sequence gives rise to an intermolecular interaction while the C-terminal domains become available for binding ligands.
    Biopolymers 06/1986; 25(5):947-58. · 2.29 Impact Factor
  • H Hörmann
    Blut 12/1985; 51(5):307-14.
  • Helmut Hörmann
    Annals of Hematology 10/1985; 51(5):307-314. · 2.40 Impact Factor
  • H Hörmann, H Richter, V Jelinić
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    ABSTRACT: Various proteolytic fragments from the central region of the fibronectin subunit chains containing the main cell-affinity site were applied in cell binding studies using peritoneal macrophages of guinea pigs. A 125I-labelled 23-kDa peptide was relatively well bound by the cells. Attachment to cells was partially inhibited by wheat germ lectin, suggesting a lectin-like site in the cell-binding domain which recognizes oligosaccharide groups with terminal N-acetylglucosamine or N-acetylneuraminic acid. Binding was inhibited by N-acetylneuraminic acid with half-maximal effect at 2 X 10(-3) M. Other inhibitors were a sialic acid rich ganglioside preparation and fetuin, a sialic acid-containing glycoprotein. In contrast to the 23-kDa peptide a 125I-labelled 125-kDa fragment was only weakly bound, although it included the sequence of the 23-kDa peptide on its C-terminus. The residual binding was weakly inhibited by low concentrations of wheat germ lectin and was remarkably improved by higher concentrations. The behavior of the peptide was explained by the presence of a sialic acid-containing oligosaccharide side chain localized outside of the 23-kDa region and interacting with the lectin-like site in the cell-binding sequence. In accord with this suggestion a 95-kDa fragment representing the oligosaccharide-containing part of the 125-kDa peptide was capable of inhibiting at least partially the cell attachment of the 23-kDa piece. The results indicate a lectin-like affinity site in the cell-binding region of fibronectin which is accessible in the 23-kDa peptide, but is masked in the 125-kDa fragment and in fibronectin by a sialic acid-containing oligosaccharide moiety.
    Hoppe-Seyler's Zeitschrift für physiologische Chemie 06/1984; 365(5):517-24.
  • H Hörmann, V Jelinić, H Richter
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    ABSTRACT: A chymotrypsin-derived and 125I-labelled 125-kDa fragment of human plasma fibronectin which contained the cell binding site, was only weakly bound by peritoneal macrophages of guinea pigs and binding was not saturable. In presence of wheat germ lectin binding increased proportionally to the logarithm of the lectin concentration. Association of 125I-fragment with cells was partially prevented by non-labelled fragment indicating a saturable receptor-ligand interaction. An apparent affinity constant of about 2--4 x 10(-5) M was evaluated. A considerable fraction of the cell-bound 125I-fragment resisted removal by proteases suggesting that it was internalized. In order to investigate an influence of wheat germ lectin on the binding of 125I-fibronectin by the cells the macrophages were preincubated with the lectin followed by washing and evaluation of 125I-fibronectin binding. A simultaneous incubation of the cells with 125I-fibronectin and lectin was impractical due to partial interaction of the two proteins giving rise to some unspecific precipitates. Preincubation with wheat germ lectin considerably improved the capacity of the macrophages for binding of 125I-fibronectin. Again the binding of 125I-labelled protein could be restricted by unlabelled one. N-acetyl-glucosamine inhibited the binding of 125I-fibronectin by wheat germ lectin-treated cells if applied in the preincubation phase and more effectively, if applied in the final 125I-fibronectin binding assay. N-Acetylneuraminic acid also inhibited this step. In addition to wheat germ lectin concanavalin A was capable of generating fibronectin receptors on the cell surface. Soy bean lectin, however, was ineffective.
    Hoppe-Seyler's Zeitschrift für physiologische Chemie 09/1983; 364(8):1011-8.
  • M Seidl, H Hörmann
    Hoppe-Seyler's Zeitschrift für physiologische Chemie 02/1983; 364(1):83-92.
  • Helmut Hörmann
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    ABSTRACT: Fibronectin, previously also termed LETS-protein, is a high-molecular-weight protein (mol. w. ca. 450,000) present in the form of thin fibrils in the pericellular space of fibroblasts and other adherent cells, as well as in distinct areas of the connective tissue. A soluble form, immunologically identical and chemically at least very similar to the cell-attached protein, is found in plasma in a concentration of about 300 micrograms/ml. It is also denominated cold-insoluble globulin. The protein has affinity both to cell surfaces and to various matrix substances such as fibrin and collagen and, therefore, is capable of mediating cell attachment to these substrates. In addition, it serves as an opsonin for the phagocytosis of gelatin-containing compounds and probably is essential for the removal of soluble fibrin from the circulating blood by the reticulo-endothelial system. Bacterial cell walls are also recognized by fibronectin. A conversion of soluble fibronectin to fibrils is achieved by heparin which also enhances the binding of soluble fibronectin to cells. Heparin or, as suggested, the related heparan sulfate present on the surface of various cells, appears to function as a cofactor in the formation of pericellular fibrils. The fibronectin fibrils precipitated with heparin, compared to soluble fibronectin, show a considerably improved affinity to native collagen, especially to type III. Hyaluronic acid has an antagonistic function which, at higher concentrations, prevents the fibronectin fibrils from interacting with collagen and cell surfaces. Masking of fibronectin fibrils was also achieved by sulfated proteoglycans of cartilage. Virus-transformed fibroblasts produce less fibronectin and are less capable of maintaining surface pericellular fibrils. A reasonable explanation is that they have an elevated secretion of hyaluronic acid. The transformed cells attach only weakly to a surface and exhibit a rounded shape in contrast to healthy ones. This phenotype can be corrected to a great extent with fibronectin. It is suggested that fibronectin also influences the formation of connective tissue by accumulating collagen precursors on the surface of fibroblasts and facilitating fibrillogenesis.
    Klinische Wochenschrift 11/1982; 60(20):1265-77.
  • Helmut Hörmann
    Journal of Molecular Medicine-jmm. 01/1982; 60(20):1265-1277.
  • H Richter, M Seidl, H Hörmann
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    ABSTRACT: Resolution of a cathepsin D digest of plasma fibronectin on heparin-Sepharose yielded, in addition to non-bound and weakly retained material (Fraction I and II), various fragments which were not eluted until 0.25M NaCl (Fraction III) and 0.5M NaCl (Fraction IV) was applied. Fraction III contained predominantly a peptide of Mr 70 000 originating from the N-terminus of the fibronectin subunits as well as high molecular weight precursors yielding the Mr 70 000 peptide by further digestion. All those peptides were retained by immobilized denatured collagen, type I, indicating the presence of the known gelatin-binding domain. In addition, they contained a transamidase-sensitive site as revealed from a digest of fibronectin previously labelled with [14C]putrescine by a transamidase-mediated reaction. Plasminolysis of the fragment of Mr 70 000 resulted in two peptides of Mr 30 000 and 40 000, only the former being retained by heparin-Sepharose. Fraction IV contained a fragment of Mr 140 000 which, after reduction, dissociated into two peptides of Mr 75 000 and 65 000. Apparently, it included the disulfide bond(s) connecting the two fibronectin subunits close to their C-terminal ends. Partial digestion of the two electrophoretically separated peptide chains with protease of Staphylococcus aureus V8 yielded for each chain a number of peptides with equal electrophoretic migration rate. In addition, however, some peptides were different in the two digests. The results were consistent with an identical or homologous structure of the two peptide chains with an additional sequence in the longer chain. The latter (Mr 75 000) uniquely contained a transamidase susceptible site as demonstrated by processing of [14C]putrescine-labelled fibronectin.
    Hoppe-Seyler's Zeitschrift für physiologische Chemie 05/1981; 362(4):399-408.
  • H Hörmann, V Jelinić
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    ABSTRACT: Trypsinized peritoneal macrophages of guinea pigs moderately bound soluble native 125I-collagen, type III. The association was slightly improved by plasma fibronectin. An additional considerable enhancement was achieved by heparin or heparan sulfate in presence of fibronectin. The enhancing effect of the two N-sulfated glycosaminoglycans was observed over a broad concentration range and, in case of heparin, exhibited a maximum near 1 mg/ml. Further rise of concentration resulted in a decline of binding. The related heparan sulfate potentiated binding less effectively than heparin. Optimal efficiency was recorded with lower concentrations. Hyaluronic acid, in higher concentrations, suppressed the enhancing effect of heparin or heparan sulfate on collagen binding. The improved attachment of native collagen, type III, to cells in presence of fibronectin and N-sulfated glycosaminoglycans was explained by a heparin/heparan sulfate induced conversion of soluble fibronectin to fibrils which showed an increased affinity to rod-like collagen molecules as well as to cell-surface structures. Hyaluronic acid, probably, inhibited the interaction of fibronectin fibrils with these substrates. Heparin and heparan sulfate, also in the absence of fibronectin, improved within a limited concentration range the binding of native 125I-collagen, type III, to the cells. The amount attached was, however, considerably less than in presence of fibronectin.
    Hoppe-Seyler's Zeitschrift für physiologische Chemie 02/1981; 362(1):87-94.