Longtime therapy of congenital factor XIII deficiency using factor XIII concentrate]

Factor XIII was determined by enzymatic and immunochemical methods in 3 patients with congenital factor XIII deficiency. Factor XIII activity measured by trans-glutaminase assay was below 1% of normal value in each of these cases. Immunelectrophoresis determination revealed the absence of the functionally active subunit A, whereas subunit S was only slightly diminished (30 to 50% of the normal value). Substitution with factor XIII concentrate caused a parallel increase of factor XIII activity and subunit A concentration. No uptake of factor XIII activity or of subunit. A by platelets could be demonstrated. Despite discontinuous substitution over a period of six years no antibody against factor XIII activity could be demonstrated in one patient with congenital factor XIII deficiency.     

Binding and degradation of blood coagulation factor XIII by cultured fibroblasts

The interaction of Factor XIII with cultured fibroblasts was examined using 125I-labeled protein and immunofluorescence. Platelet or plasma Factor XIII bound to confluent cell layers. Binding reached an apparent steady state after 8 h. Activation with thrombin increased the binding of both the platelet and plasma forms of the enzyme. After a 1-2 h lag, a chloroquine-inhibitable increase in trichloroacetic acid-soluble radioactivity was detected in the medium. Gel electrophoresis in sodium dodecyl sulfate indicated that approximately 16-fold more a subunit (catalytic) of 125I-plasma Factor XIII bound to the cell layer than b subunit (carrier) and that some large complexes containing Factor XIII were formed with the cell layer. Factor XIII binding increased linearly with concentrations of Factor XIII up to 230 micrograms/ml, whereas a component of the degradation of Factor XIII was saturable at about 20 micrograms/ml. Factor XIII associated with cell layers was catalytically active since it could cross-link fibronectin. By immunofluorescence the a subunit of Factor XIII was localized to fibronectin-containing extracellular fibrils and, in the presence of chloroquine, to intracellular granules. These results indicate that the a subunit of Factor XIII binds to the fibroblast extracellular matrix and matrix assembly sites, where it remains active, and to a putative cell-surface receptor which mediates its internalization and degradation.     

Tb(III)-ion-binding-induced conformational changes in platelet factor XIII.

Ca(II) ions are crucial during proteolytic conversion of Factor XIII zymogen into the active enzyme Factor XIIIa. Factor XIII proteolyzed by thrombin or trypsin in the presence of 5 mM-EDTA resulted in rapid inactivation of transglutaminase activity. Factor XIIIa formed by thrombin or trypsin in the presence of 40 microM-Tb(III) ions, however, was indistinguishable from Factor XIIIa formed in the presence of 2-5 mM-Ca(II) ions with respect to molecular mass and transglutaminase activity. Thrombin treatment of Factor XIII in the presence of 1-5 microM-Tb(III) ions resulted in three fragments (76 kDa, 51 kDa and 19 kDa) with simultaneous loss of transglutaminase activity. Tb(III) ions at concentrations greater than 40 microM made platelet Factor XIII resistant to proteolysis by either thrombin or trypsin. Other lanthanide(III) ions [Ln(III) ions] tested [Ce(III), La(III) and Gd(III) ions] functioned similarly to Tb(III) ions during proteolytic activation of Factor XIII. Ln(III) ions (10-100 microM) were unable to replace the Ca(II) ions required for transglutaminase activity of Factor XIIIa. Tb(III) ions also inhibited in a non-competitive manner the transglutaminase activity of Factor XIIIa (Ki 71 microM) even when measured in the presence of 200-fold molar excess of Ca(II) ions. Factor XIII selectively bound to a Tb(III)-chelate affinity column, and could not be eluted by 100 mM-CaCl2. Binding of Tb(III) ions to Factor XIII was demonstrated by fluorescence emission due to Forster energy transfer. A 10(4)-fold molar excess of CaCl2, but not NaCl, partially quenched Tb(III) fluorescence. Low concentrations (5-20 microM) of Tb(III) ions also inhibited the binding of Factor XIII to des-A-fibrinogen by about 43%, whereas higher concentrations (40-100 microM) promoted binding. Conformational changes in Factor XIII consequent to the binding of Tb(III) ions could be responsible for the observed effects on protein structure and function.     

b-chains prevent the proteolytic inactivation of the a-chains of plasma factor XIII

While the transglutaminase activity is associated exclusively with the thrombin-cleaved a chains of plasma Factor XIII, there is little information regarding the role of the b-chains. The present investigations were undertaken to clarify the role of the b-chains during proteolytic activation of plasma factor XIII a-chains. The a-chains of platelet Factor XIII (a2) were extremely sensitive to alpha-thrombin proteolysis, especially in the presence of 5 mM EDTA, resulting in two major fragments with molecular masses 51 +/- 3 kDa and 19 +/- 4 kDa. Furthermore, fibrin enhanced the alpha-thrombin proteolysis of thrombin-cleaved platelet Factor XIII a-chains in presence of CaCl2 or EDTA, resulting in several peptide fragments with molecular masses from 51 +/- 3 kDa to 14 +/- 4 kDa. By contrast, thrombin-cleaved a-chains of plasma Factor XIII (a2b2) were not further degraded by alpha-thrombin in presence of 5 mM EDTA. Even in the combined presence of 5 mM EDTA and 0.1 mg/ml fibrin, alpha-thrombin proteolysis of plasma Factor XIIIa was limited to the formation of a 76 kDa fragment (= Factor XIIIa), a 51 +/- 3 kDa fragment and trace amounts of a 14 +/- 4 kDa species. Platelet Factor XIII proteolyzed by 500 nM alpha-thrombin in presence of 5 mM EDTA expressed less than 20% of enzymatic activity obtained when platelet Factor XIII was activated in presence of 5 mM CaCl2. In contrast, plasma Factor XIII activated by 500 nM apha-thrombin in presence of 5 mM EDTA expressed nearly 65% of original transglutaminase activity. Likewise, when plasma Factor XIII was proteolyzed by 100-1000 nM gamma-thrombin in presence of 5 mM CaCl2 or 5 mM EDTA, maximal transglutaminase activity was observed. However, when platelet Factor XIII was similarly treated with gamma-thrombin in presence of 5 mM EDTA, only one-half the original transglutaminase activity was obtained. The b-chains thus appear to mimic the function of Ca2+ in preserving transglutaminase activity of thrombin-cleaved a-chains. The b-chains of plasma Factor XIII were not degraded by either alpha- or gamma-thrombin treatment, in presence of 5 mM EDTA or 5 mM CaCl2. Both platelet and plasma Factor XIII a-chains were degraded by trypsin to fragments with molecular masses of 51 +/- 3 kDa and 19 +/- 4 kDa in presence of 5 mM CaCl2 and to fragments with molecular masses of 19 +/- 4 kDa and lower, in presence of 5 mM EDTA.(ABSTRACT TRUNCATED AT 400 WORDS)     

Examining thrombin hydrolysis of the factor XIII activation peptide segment leads to a proposal for explaining the cardioprotective effects observed with the factor XIII V34L mutation

In the blood coagulation cascade, thrombin cleaves fibrinopeptides A and B from fibrinogen revealing sites for fibrin polymerization that lead to insoluble clot formation. Factor XIII stabilizes this clot by catalyzing the formation of intermolecular cross-links in the fibrin network. Thrombin activates the Factor XIII a(2) dimer by cleaving the Factor XIII activation peptide segment at the Arg(37)-Gly(38) peptide bond. Using a high performance liquid chromatography assay, the kinetic constants K(m), k(cat), and k(cat)/K(m) were determined for thrombin hydrolysis of fibrinogen Aalpha-(7-20), Factor XIII activation peptide-(28-41), and Factor XIII activation peptide-(28-41) with a Val(34) to Leu substitution. This Val to Leu mutation has been correlated with protection from myocardial infarction. In the absence of fibrin, the Factor XIII activation peptide-(28-41) exhibits a 10-fold lower k(cat)/K(m) value than fibrinogen Aalpha-(7-20). With the Factor XIII V34L mutation, decreases in K(m) and increases in k(cat) produce a 6-fold increase in k(cat)/K(m) relative to the wild-type Factor XIII sequence. A review of the x-ray crystal structures of known substrates and inhibitors of thrombin leads to a hypothesis that the new Leu generates a peptide with more extensive interactions with the surface of thrombin. As a result, the Factor XIII V34L is proposed to be susceptible to wasteful conversion of zymogen to activated enzyme. Premature depletion may provide cardioprotective effects.     

Characterization of the fibrin polymer structure that accelerates thrombin cleavage of plasma factor XIII

The effect of plasmin-derived fibrin(ogen) degradation products on alpha-thrombin cleavage of plasma Factor XIII was studied to identify the fibrin polymer structure that promotes Factor XIIIa formation. Fibrin polymers derived from fibrinogen and Fragment X enhanced the rate of thrombin cleavage of plasma Factor XIII in plasma or buffered solutions. The concentrations of fibrinogen and Fragment X that promoted half-maximal rates of Factor XIIIa formation were 5 and 40 micrograms/ml, respectively. Fragments Y, D, E, D-dimer, and photooxidized fibrinogen did not enhance thrombin cleavage of Factor XIII. Although purified Fragment D1 inhibited fibrin gelation, the soluble protofibrils promoted thrombin activation of Factor XIII. Noncrosslinked fibrin fibers failed to enhance thrombin cleavage of Factor XIII. In conclusion, soluble fibrin oligomers function to promote thrombin cleavage of plasma Factor XIII during blood clotting.     

Change in factor XIII activity in plasma during thrombocyte aggregation]

On addition to rat plasma, platelet-enriched, of active factor XIII (XIIIa), there occurred along with increase in the aggregation induced by ADP, a reduction in the activity of factor XIII in the plasma. Addition to the plasma of inactive factor XIII failed to influence either the degree of aggregation, or the change in the activity of factor XIII in the plasma in comparison with control samples. In platelet aggregation induced by thrombin, addition of factor XIII was accompanied by a marked fall of its activity in the plasma. In comparison with control, the extent of aggregation in this case decreased. The observed differences in the character of aggregation coursing in the presence of factor XIIIa when different aggregating agents (ADP and thrombin) were used were apparently due to the interaction of active factor XIII with thrombin added to the plasma in the capacity of an aggregating agent.     

Successful pregnancy in a woman with congenital factor XIII deficiency treated with substitutive therapy. Report of a second case

A syndrome of marked fetal wastage is associated with congenital factor XIII deficiency in adult women. A previously unreported case of a woman with factor XIII deficiency is described, in which substitutive treatment with normal plasma or placental factor XIII concentrate permitted two normal pregnancies. Factor XIII activity was maintained above 1-2% with intermittent infusion of 300 ml to 450 ml of plasma every 14 days or of 500 units of concentrate every 21 days. This case confirms the only other case so far reported in which factor XIII substitutive therapy was able to permit a normal pregnancy in a woman with factor XIII deficiency and seems to suggest factor XIII to be involved in the process of annidation.     

Structural features associated with the binding of glutamine-containing peptides to Factor XIII

Activated Factor XIII a2 catalyzes the formation of intermolecular gamma-glutamyl- epsilon -lysyl cross-links in the fibrin network. Solution NMR studies were carried out to characterize, the structural features associated with the binding of glutamine-containing peptides to Factor XIII. A coupled uv/vis kinetic assay demonstrated that K9 peptide (1-10), alpha2-antiplasmin (1-15), and alpha2-antiplasmin (1-15 Q4N) all function as glutamine-containing substrates for activated Factor XIII a2. 2D TOCSY spectra of the peptides exhibit upfield chemical shifts for the glutamine protons in the presence of Factor XIII. These results indicate that the reactive peptide glutamines are encountering a distinctive environment within the Factor XIII active site. 1D proton line-broadening and 2D transferred-NOESY studies reveal that the glutamines and residues located C-terminally come in direct contact with the enzyme and adopt an extended conformation. Substrates with sequences similar to alpha2-antiplasmin (1-15) are proposed to bind both at the catalytic site and at a neighboring apolar region.     

The binding of divalent metal ions to platelet factor XIII modulates its proteolysis by trypsin and thrombin

We investigated the effect of divalent metal ions on the proteolytic cleavage and activation of platelet Factor XIII by thrombin and trypsin. In the absence of metal ions (5 mM EDTA), trypsin and thrombin rapidly degraded platelet Factor XIII (80 kDa) to low-molecular-mass peptides (50-19 kDa) with simultaneous loss of transglutaminase activity. Divalent metal ions protected Factor XIII from proteolytic inactivation with an order of efficacy of Ca2+ greater than Zn2+ greater than Mg2+ greater than Mn2+. Calcium (2 mM) increased by 10- to 1000-fold the trypsin and thrombin concentrations required to degrade Factor XIII to a 19-kDa peptide. Factor XIIIa formed by thrombin in the presence of 5 mM EDTA had one-half the specific activity of Factor XIIIa formed in the presence of calcium. Factor XIII was cleaved by trypsin in the presence of 5 mM Ca2+ to a 51 +/- 3-kDa fragment that had 60% of the original Factor XIIIa activity. A similar tryptic peptide formed in the presence of 5 mM EDTA did not have transglutaminase activity. In the presence of 5 mM Mg2+, thrombin cleaved Factor XIII to a major 51 +/- 3-kDa fragment that had 60% of the Factor XIIIa activity. Mn2+ (0.1-5 mM) limited trypsin and thrombin proteolysis. The resulting digest containing a population of Factor XIII fragments (50-14 kDa) expressed 50-60% transglutaminase activity of Factor XIIIa. Factor XIII was fully activated by both trypsin and thrombin in the presence of 5 mM Zn2+, resulting in two fragments of 76 and 72 kDa. We conclude that the binding of divalent metal ions to platelet Factor XIII induces conformational changes in the protein that alter its susceptibility to proteolysis and influence the expression of transglutaminase activity.     

Amino acid sequence of the a subunit of human factor XIII

Factor XIII is a plasma protein that plays an important role in the final stages of blood coagulation and fibrinolysis. The complete amino acid sequence of the a subunit of human factor XIII was determined by a combination of cDNA cloning and amino acid sequence analysis. A lambda gtll cDNA library prepared from human placenta mRNA was screened with an affinity-purified antibody against the a subunit of human factor XIII and then with a synthetic oligonucleotide probe that coded for a portion of the amino acid sequence present in the activation peptide of the a subunit. Six positive clones were identified and shown to code for the a subunit of factor XIII by DNA sequence analysis. A total of 3831 base pairs was determined by sequencing six overlapping cDNA clones. This DNA sequence contains a 5' noncoding region or a region coding for a portion of a pro-piece or leader sequence, the mature protein (731 amino acids), a stop codon (TGA), a 3' noncoding region (1535 nucleotides), and a poly(A) tail (10 nucleotides). When the a subunit of human factor XIII was digested with cyanogen bromide, 11 peptides were isolated by gel filtration and reverse-phase HPLC. Amino acid sequence analyses of these peptides were performed with an automated sequenator, and 363 amino acid residues were identified. These amino acid sequences were in complete agreement with those predicted from the cDNA. The a subunit of factor XIII contained the active site sequence of Tyr-Gly-Gln-Cys-Trp, which is identical with that of tissue transglutaminase.(ABSTRACT TRUNCATED AT 250 WORDS)     

Evaluating the roles of thrombin and calcium in the activation of coagulation factor XIII using H/D exchange and MALDI-TOF MS

Factor XIII catalyzes the formation of isopeptide bonds between noncovalently associated fibrin monomers in the final stages of the blood coagulation cascade. This results in a rigid, covalently linked network that is much more resistant to proteolytic degradation. Calcium ion is critical to this process, and its continued presence after activation aids in maintenance of Factor XIII activity. Hydrogen/deuterium exchange experiments were conducted on recombinant Factor XIII a(2) using matrix-assisted laser desorption ionization time-of-flight mass spectrometry. The method revealed changes in the structure of Factor XIII a(2) localized to different areas of the protein that were related to the manner in which the enzyme was activated and the calcium environment in which it was maintained. A possible substrate recognition region in the catalytic core (220-230) shows an increase in deuteration upon activation. The degree of deuteration varies depending on the calcium environment in which the active enzyme is maintained. A portion of the beta-sandwich domain (98-104) exhibits a decrease in deuteration upon activation by exposure to calcium alone. A third change occurs in the beta-barrel 1 domain of the protein, a portion of which (526-546) shows a decrease in deuteration upon activation by calcium exposure, but almost none at all when the enzyme is activated by thrombin. The pattern of observed changes reveals individual contributions of calcium and thrombin to activating the enzyme toward substrate binding and exposure of the active site.     

Isolation of a fibrin-binding fragment from blood coagulation factor XIII capable of cross-linking fibrin(ogen).

Purified platelet Factor XIII was radioiodinated and then partially degraded by thrombin or trypsin, and a fibrin-binding fragment was identified by autoradiography and immunoblotting following separation by SDS/polyacrylamide-gel electrophoresis. Limited proteolysis of 125I-Factor XIII by thrombin or trypsin produced an 125I-51 kDa fragment and an unlabelled 19 kDa fragment. The 51 kDa fragment was purified by h.p.l.c. on a TSK-125 gel-filtration column. Partial amino acid sequence analysis of the 51 kDa fragment indicated that it was similar in sequence to the Gly38-Lys513 segment in placental Factor XIII a-chain. More than 70% of the 51 kDa fragment bound to fibrin, whereas the 19 kDa fragment did not bind. The active site was localized to the 51 kDa fragment since this fragment expressed transglutaminase activity, cross-linked fibrin and fibrinogen and incorporated iodo[14C]acetamide into the active-site cysteine residue. Isolation of a fibrin-binding fragment expressing transglutaminase activity demonstrates that each a-chain of the dimeric Factor XIIIa could function independently to cross-link fibrin. The fibrin-binding site could play an important role in localizing Factor XIIIa to the fibrin clot.     

Cross-linking of salmon fibrinogen and fibrin by factor XIII and transglutaminase

Unlike mammalian species, salmon plasma contains 2 cross-linking enzyme systems: Factor XIII and a transglutaminase which appears to be similar in its action to that described by Folk and Chung. Also, salmon plasma contains an exceedingly active protease which possesses the ability to rapidly destroy fibrinogen clottability even when the plasma is stored at a temperature of ?20°C.     

Separation of human plasma fibrin stabilizing factor (factor XIII) from fibrinogen by fibrinogen polymer formation

Human plasma fibrin stabilizing factor (factor XIII) may be separated from fibrinogen through reversible fibrinogen polymer formation at pH 6.6, gamma/2 0.3, 0 degrees C, and subsequent Bio-Gel A 1.5m filtration. Factor XIII activity is eluted after the monomer fibrinogen peak. Polymer fractions from eight preparations, processed in duplicate, contain a mean 0.002 units factor XIII per mg fibrinogen, or about 0.7% the factor XIII content of standard plasma. Factor XIII-free fibrinogen polymers are easily dissociated (greater than 98%) to the monomer form by incubation at 37 degrees C, 18 hours. The fibrinogen preparations utilized were devoid of plasma fibronectin; thus these studies also show that reversible human fibrinogen polymer formation occurs in its absence.     

Calcium-induced dissociation of human plasma factor XIII and the appearance of catalytic activity

1. The Ca(2+) dependence of the activity of plasma Factor XIII(a) was studied by using the continuous assay based on the incorporation of dansylcadaverine into dephosphorylated acetylated beta-casein (beta-substrate). The K(m) for Ca(2+) is about 0.170mm. 2. At low concentrations of Ca(2+) there was a lag in attaining the steady-state rate. The size of the lag was decreased and eventually abolished if the enzyme was preincubated with a high concentration of Ca(2+) before assay. The concentration of Ca(2+) required to decrease the lag phase by 50% in 10min depended on the protein concentration: at 0.87mg of protein/ml it required 17mm-Ca(2+) and at 0.44mg/ml it needed 10mm-Ca(2+). 3. The concentrations of Ca(2+) required either to abolish the lag phase in the appearance of enzyme activity or to activate the essential thiol for reaction with 5,5'-dithiobis-(2-nitrobenzoate) in 10min incubation were similar at the same protein concentration. This indicated that Ca(2+) induces a conformation change that is responsible for both phenomena. A model is proposed that links this conformation change to the dissociation of the tetrameric enzyme. 4. This was supported by the observation that the addition of excess of b chains to the Factor XIII(a) (a'(2)b(2)) increased the concentration of Ca(2+) required to expose the reactive thiol, and inhibited the Ca(2+)-dependent aggregation of a' chains. 5. Platelet Factor XIII(a) (a'(2)) was inhibited by 5,5'-dithiobis-(2-nitrobenzoate) in the absence of Ca(2+), and no lag phases were observed in attaining the steady-state rate at low Ca(2+) concentrations, thus confirming the model for the activation of the plasma enzyme. 6. The Ca(2+) dependence of platelet Factor XIII(a) indicated that Ca(2+) has an additional role in the enzyme mechanism of the plasma enzyme, perhaps being involved in substrate binding. 7. The dependence of the stability of plasma Factor XIII(a) on Ca(2+) and protein concentration indicates that the decay in activity is related to the tetramer dissociation. 8. beta-Substrate decreased the Ca(2+) concentration required for (1) abolition of the lag phase and (2) enzyme inhibition by thiol reagents. The effect on the former is greater than on the latter. 9. The role of the b chains of the plasma Factor and the evolutionary significance of the plasma and platelet Factors are considered.     

Amino acid sequence of the peptide released from bovine factor XIII following activation by thrombin

The amino acid sequence of the peptide released during the conversion of bovine Factor XIII to the active enzyme by thrombin was determined. It contains N-terminal N-acetylserine and a total of 37 residues. The bovine peptide differs from the corresponding human peptide. There are 5 amino acid replacements and one deletion in the human peptide.     

On the activation of bovine plasma factor XIII. Amino acid sequence of the peptide released by thrombin and the terminal residues of the subunit polypeptides.

A blood coagulation factor, Factor XIII, was highly purified from bovine fresh plasma by a method similar to those used for human plasma Factor XIII. The isolated Factor XIII consisted of two subunit polypeptides, a and b chains, with molecular weights of 79,000 +/- 2,000 and 75,000 +/- 2,000, respectively. In the conversion of Factor XIII to the active enzyme, Factor XIIIa, by bovine thrombin [EC 3.4.21.5], a peptide was liberated. This peptide, designated tentatively as "activation peptide," was isolated by gel-filtration on a Sephadex G-75 column. It contained a total of 37 amino acid residues with a masked N-terminal residue and C-terminal arginine. The whole amino acid sequence of "Activation peptide" was established by the dansyl-Edman method and standard enzymatic techniques, and the masked N-terminal residue was identified as N-acetylserine by using a rat liver acylamino acid-releasing enzyme. This enzyme specifically cleaved the N-acetylserylglutamyl peptide bond serine and the remaining peptide, which was now reactive to 1-dimethylamino-naphthalene-5-sulfonyl chloride. A comparison of the sequences of human and bovine "Activation peptide" revealed five amino acids replacements, Ser-3 to Thr; Gly-5 to Arg; Ile-14 to Val; Thr-18 to Asn, and Pro-26 to Leu. Another difference was the deletion of Leu-34 in the human peptide. Adsorption chromatography on a hydroxylapatite column in the presence of 0.1% sodium dodecyl sulfate was developed as a preparative procedure for the resolution of the two subunit polypeptides, a or a' chain and b chain, constituting the protein molecule of Factor XIII or Factor XIIIa. End group analyses on the isolated pure chains revealed that the structural change of Factor XIII during activation with thrombin occurs only in the N-terminal portion of the a chain, not in the N-terminal end of the b chain or in the C-terminal ends of the a and b chains. From these results, it was concluded that the activation of bovine plasma Factor XIII by thrombin must be accompanied by a limited proteolysis of the arginyl-glycyl bond located in the N-terminal region of the a chain, liberating the "Activation peptide." The possibility of activating Factor XII with other porteinases was examined using Factor Xa [EC 3.4.21.6], Factor XIIa, kallikreins [EC 3.4.21.8], urokinase [EC 3.4.99.26], trypsin [EC 3.4.21.4], ficin [EC 3.4.22.3], papain [EC 3.4.22.2], and bromelain [EC 3.4.22.4]. Among these enzymes, only bromelain and trypsin showed clear activating effects.     

An equilibrium study of metal ion binding to human plasma coagulation factor XIII.

1. The binding of Ca2+ to plasma coagulation Factor XIII from man and from cow caused a small decrease in the intrinsic fluorescence of the protein with a dissociation constant of 0.1 mM. A similar decrease was observed with the thrombin-activated Factors (Factors XIIa). The decrease in protein fluorescence was also caused by both Ni2+ and Mn2+ but not by Mg2+. 2. 45Ca2+ binding was directly demonstrated by equilibrium dialysis. Ca2+ at 0.2 mM bound to Factor XIII (a2b2) and Factor XIIIa (a'2b2) but not to isolated b2-protein. A tight-binding site for Ca2+ is associated with the a-subunits. 3. The Ca2+ essential for the enzyme activity of Factor XIII from man, pig and cow can be replaced by Ni2+, Cu2+, La3+, Mn2+, Fe3+, Y3+, Co2+, Sr2+ or Tb3+, but not by Mg2+.     

Interactions of factor XIII with fibrin as substrate and cofactor.

Factor XIIIa (a2') is a homodimeric transglutaminase that is formed via limited alpha-thrombin-catalyzed proteolysis of the platelet (a2) or plasma (a2b2) factor XIII zymogen in a reaction that results in proteolytic removal of a 37-aminoacyl residue peptide from the N-terminus of the a chains and exposure of the active-site thiol group in the resulting a' chains of factor XIIIa. In this study, we characterized interactions of factor XIII and factor XIIIa with fibrin, a natural substrate for factor XIIIa and a cofactor for the alpha-thrombin-catalyzed activation of plasma factor XIII. The carbamylmethyl derivatives of the active-site thiol group of platelet factor XIII (CMa2) and factor XIIIa (CMa2') were prepared, and their interactions with fibrin were measured. The enzyme-like derivative (CMa2') which contained nicked a' chains bound more tightly to fibrin (Kd = 2.1 microM) than did CMa2 (Kd = 14 microM), the platelet zymogen-like derivative with intact a chains, but the binding of each was weaker than the binding of plasma factor XIII zymogen (a2b2) to fibrin (Kd = 0.20 microM) under the same conditions. Saturation of fibrin with plasma factor XIII zymogen (a2b2) did not affect the binding of CMa2' to fibrin, suggesting that the plasma factor XIII zymogen (a2b2) and the active-site-modified form of factor XIIIa (CMa2') bind to separate, noninteracting sites of fibrin.(ABSTRACT TRUNCATED AT 250 WORDS)