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PubMed1998

Structural determinants of the bifunctional corn Hageman factor inhibitor: x-ray crystal structure at 1.95 A resolution.

Corn Hageman factor inhibitor (CHFI) is a bifunctional 127 residue, 13.6 kDa protein isolated from corn seeds. It inhibits mammalian trypsin and Factor XIIa (Hageman Factor) of the contact pathway of coagulation as well as alpha-amylases from several insect species. Among the plasma proteinases, CHFI specifically inhibits Factor XIIa without affecting the activity of other coagulation proteinases. We have isolated CHFI from corn and determined the crystallographic structure at 1.95 A resolution. Additionally, we have solved the structure of the recombinant protein produced in Escherichia coli at 2.2 A resolution. The two proteins are essentially identical. The proteinase binding loop is in the canonical conformation for proteinase inhibitors. In an effort to understand alpha-amylase inhibition by members of the family of 25 cereal trypsin/alpha-amylase inhibitors, we have made three-dimensional models of several proteins in the family based on the CHFI coordinates and the coordinates determined for wheat alpha-amylase inhibitor 0.19 [Oda, Y., Matsunaga, T., Fukuyama, K., Miyazaki, T., and Morimoto, T. (1997) Biochemistry 36, 13503-13511]. From an analysis of the models and a structure-based sequence analysis, we propose a testable hypothesis for the regions of these proteins which bind alpha-amylase. In the course of the investigations, we have found that the cereal trypsin/alpha-amylase inhibitor family is evolutionarily related to the family of nonspecific lipid-transfer proteins of plants. This is a new addition to the group which now consists of the trypsin/alpha-amylase inhibitors, 2S seed storage albumins, and the lipid-transfer family. Apparently, the four-helix conformation has been a successful vehicle in plant evolution for providing protection from predators, food for the embryo, and lipid transfer.

Amino Acid Sequence↗
PubMed1983

Activation of factor XII by dextran sulfate: the basis for an assay of factor XII.

A system was developed for studying the activation of factor XII (Hageman factor) in the presence of dextran sulfate (DS). Salient features of the system included low ionic strength (0.08), low concentration of factor XII (approximately 1/10,000 that in normal plasma), and an excess of exogenous prekallikrein (PK). In this system, factor XII was rapidly converted to the 80,000 molecular weight (mol wt) form of factor XIIa (alpha-factor-XIIa). Once formed, the factor XIIa converted PK to kallikrein at a rate that was proportional to the amount of factor XII originally present in the incubation mixture. This system was used to construct a simple sensitive assay for factor XII in plasma and other biologic samples. The kallikrein produced was measured spectrophotometrically with the chromogenic substrate (H-D-Pro-Phe-Arg-p-nitroanilide (S-2302). This assay was shown to be independent of the high molecular weight kininogen and the PK content of the sample being analyzed. The measurements obtained were consistent with fundamental enzymologic principles and, if desired, could be processed with a simple calculator program to achieve linear standard curves. When applied to the quantitation of factor XII in plasma, the assay yielded values in close agreement with those determined by coagulant assay or by radial immunodiffusion.

Dextran Sulfate↗
PubMed1994

Inhibition of serine proteases of the blood coagulation system by squash family protease inhibitors.

Squash family inhibitors are the smallest protein serine protease inhibitors, being composed of approximately 30 amino acid residues. We isolated 8 squash family inhibitors from the seeds of bitter gourd, squash, gourd and luffa and examined their effect on serine proteases of the blood coagulation system. Five of them prolonged the activated partial thromboplastin time of human plasma to various extents, but three did not. Only Momordica charantia (bitter gourd) trypsin inhibitor-II prolonged the prothrombin time of human plasma. All inhibitors inhibited the amidolytic activities of factor XIIa, plasma kallikrein, factor Xa, but did not inhibit significantly those of factor XIa, factor IXa, factor VIIa, and thrombin. Ki values for factor XIIa, plasma kallikrein, and factor Xa were in the order of 10(-6)-10(-9), 10(-4)-10(-5), and 10(-4)-10(-6)M, respectively. The prolongation of the activated partial thromboplastin time by inhibitors appeared to correspond to their inhibitory potencies for factor XIIa. Momordica charantia trypsin inhibitor-II, which has the strongest inhibitory potency toward the amidolytic activity of factor Xa, with a Ki value 10-100 times smaller than those of other inhibitors, inhibited the activation of factor X by factor VIIa-tissue factor complex or factor IXa, while others did not.

Amino Acid Sequence↗
PubMed1994

Proteolytic activation of single-chain precursor macrophage-stimulating protein by nerve growth factor-gamma and epidermal growth factor-binding protein, members of the kallikrein family.

Promacrophage-stimulating protein (MSP) is an 80-kDa protein that acquires biological activity after cleavage at an Arg-Val bond to a disulfide-linked alpha beta heterodimer by serine proteases of the intrinsic coagulation cascade. These proteases, which include serum kallikrein, factor XIIa and factor XIa, are members of the trypsin family of serine proteases. We now report that two other members of the family, nerve growth factor-gamma (NGF-gamma) and epidermal growth factor-binding protein (EGF-BP), cleave and activate pro-MSP to the disulfide-linked alpha beta heterodimer. Cleavage of 1.5 nM pro-MSP by 1 nM NGF-gamma or EGF-BP at 37 degrees C was almost complete within 30 min. These concentrations of enzyme are about 2 orders of magnitude less than is required for cleavage by serum kallikrein or factor XIIa. Cleavage of pro-MSP to MSP was associated with a conformational change in the protein, because the cleaved product, but not pro-MSP, was detected by a sandwich enzyme-linked immunoassay. Cleavage caused the appearance of biological activity, as measured by chemotactic activity of MSP for resident peritoneal macrophages, by MSP-induced macrophage shape change, and by stimulation of macrophage ingestion of C3bi-coated erythrocytes. These findings suggest the possibility of cooperative interactions between NGF-gamma or EGF-BP and pro-MSP in inflammation and wound healing.

Animals↗
PubMed1994

Proteolytic conversion of single chain precursor macrophage-stimulating protein to a biologically active heterodimer by contact enzymes of the coagulation cascade.

Human serum macrophage stimulating protein (MSP) is a disulfide-linked heterodimer that induces motile and phagocytic activity of mouse resident peritoneal macrophages. It is a member of the family of kringle proteins, which typically exist in extracellular fluid as single chain precursors that are activated by proteolytic cleavage. In this work, we expressed [35S]cysteine-labeled recombinant pro-MSP in MSP cDNA-transfected Chinese hamster ovary cells and studied proteolytic processing of pro-MSP and the requirement of cleavage for biological activity. In media containing heat-inactivated fetal bovine serum, the protein was secreted as single chain pro-MSP, which was cleaved over a period of hours to the mature heterodimer. Cleavage was prevented by serine protease inhibitors such as leupeptin or aprotinin; it did not occur if cells were cultured in serum-free medium. Nanomolar concentrations of coagulation proteases kallikrein, factor XIIa or factor XIa cleaved pro-MSP to MSP within 30 min. Pro-MSP had no biological activity. After cleavage by kallikrein, biological activity was quantitatively comparable to that of natural MSP isolated from human plasma. These results support our hypothesis that MSP circulates as the biologically inactive precursor and can be activated by enzymes of the intrinsic coagulation cascade.

Animals↗
PubMed1987

Structural and functional characterization of factor XII.

In this article we have reviewed the current knowledge regarding the involvement of Factor XII in contact activation. Clearly in the past decade an overwhelming amount of data and hypotheses have been published regarding the central role of this zymogen in the initiation and further propagation of contact activation reactions. Therefore we feel that it will be helpful to conclude this article with a figure that summarizes those interactions and reactions that are generally believed to reflect the major molecular events occurring during surface-dependent contact activation. The contact factors are capable of very efficient interation with each other, provided a suitable negatively charged surface is present. Such surfaces are thought to stimulate the interactions between the contact factors through binding of the proteins and thus bringing the proteins together. Factor XII readily binds to the negatively charged surface, but for the binding of prekallikrein and Factor XI, the cofactor HMW kininogen is likely to be necessary. Bound at the surface, the zymogens Factor XII and prekallikrein are thought to be involved in a so-called reciprocal activation mechanism in which Factor XIIa activates prekallikrein to kallikrein, which in turn converts Factor XII to Factor XIIa. The formation of Factor XIIa is further promoted by the fact that surface-bound Factor XII is likely more susceptible to proteolytic cleavage and by the fact that the activated Factor XIIa is capable of auto-activating its own zymogen Factor XII. However, the latter effect, although undoubtedly contributing to the formation of Factor XIIa at the surface, seems to be of less importance than the reciprocal activation mechanism. This is underscored by the fact that Factor XII activation is rather slow in prekallikrein-deficient plasma. Surface-bound Factor XIIa is then responsible for the activation of Factor XI to Factor XIa, thereby propagating the initial trigger. Presumably, Factor XIa must leave the surface in order to be able to become involved in the activation of blood coagulation Factor IX.

Amino Acid Sequence↗
PubMed1988

Modulation of the human monocyte binding site for monomeric immunoglobulin G by activated Hageman factor.

Macrophage Fc gamma receptors play a significant role in inflammation and host defense. One monocyte/macrophage Fc gamma receptor, Fc gamma RI, the binding site for monomeric IgG, appears to be especially responsive to modulatory signals by hormones and mediators. Since Factor XIIa is generated during inflammation, we studied the effect of XIIa on Fc gamma RI. Factor XIIa, in a concentration-dependent manner (0.01-0.19 microM), reduced the number of monocyte binding sites for monomeric IgG up to 80% without altering the affinity of binding. Its precursor, Factor XII, and the low molecular weight fragment of XIIa, lacking most of the heavy chain region, did not reduce the expression of Fc gamma RI. Neither corn trypsin inhibitor (36 microM) nor diisopropylfluorophosphate (3.6 mM) diminished the effect of Factor XIIa on Fc gamma RI, although each completely inhibited the coagulant and amidolytic activity contained on the light chain of Factor XIIa. Protein synthesis was not a requirement for this effect of Factor XIIa, nor was internalization of Fc gamma RI necessary. In contrast to similar concentrations of IgG, Factor XIIa failed to displace significantly monomeric IgG from the monocyte surface, suggesting that Factor XIIa does not directly compete for Fc gamma RI. The data suggest that the heavy chain of XIIa, which contains domains that may have cell hormone activity, also contains a domain that regulates Fc gamma RI on monocytes. In addition to other hormones and mediators, Factor XIIa may serve a regulatory function in modulating Fc gamma receptor expression during inflammation.

Binding Sites↗
PubMed1991

Zinc ions promote the binding of factor XII/factor XIIA to acidic phospholipids but have no effect on the binding of high-Mr kininogen.

Binding of high-Mr kininogen and factor XII/factor XIIa to phospholipids coated on to polystyrene microtiter plates was investigated by ELISA. Both high-Mr kininogen and factor XII/factor XIIa bound specifically to the phospholipid surface. Binding was observed to negatively charged phospholipids only. The binding of high-Mr kininogen was not affected by the presence of zinc ions. At a surface concentration of 20% phosphatidylinositol phosphate in phosphatidylcholine a dissociation constant (kD) of 10 nM for the binding of high-Mr kininogen was calculated. The amount of bound purified alpha-factor XIIa could be increased 4-5-fold in the presence of zinc ions. The lowest zinc ion concentration giving maximal binding was 0.1 mM. The binding of alpha-factor XIIa was inhibited by high-Mr kininogen. Independent of the presence of zinc ions or high-Mr kininogen, a kD of 7.9 nM was calculated for alpha-factor XIIa binding. The binding of prekallikrein was dependent upon the presence and the concentration of high-Mr kininogen. In plasma containing aprotinin, the binding of high-Mr kininogen was apparently inhibited in the presence of zinc ions, which was a prerequisite for the binding of factor XII. This apparently inhibitory effect of zinc ions on the binding of high-Mr kininogen was probably due to the increased binding of factor XII, which displaced high-Mr kininogen.

Binding Sites↗
PubMed2006

Impaired factor XIIa-dependent activation of fibrinolysis in treated antiphospholipid syndrome gestations developing late-pregnancy complications.

OBJECTIVE: The objective of the study was to investigate the potential role of impaired factor XII-dependent activation of fibrinolysis in treated antiphospholipid syndrome gestations developing late-pregnancy complications. STUDY DESIGN: This was a prospective study in a third-level teaching hospital, including 75 patients: 25 pregnant patients having the antiphospholipid syndrome and carrying their pregnancies until 26 weeks' gestation or later (group 1); 25 pregnant patients having normal term pregnancies and delivery and no previous miscarriage (group 2); and 25 pregnant patients being diagnosed as having severe pre-eclampsia and/or intrauterine growth restriction but testing negative for antiphospholipid antibodies (group 3). Hemostatic evaluation was carried out from patients in groups 1 and 2 between 6 and 10 weeks, between 18 and 22 weeks, and between 28 and 32 weeks' gestation. Patients in group 3 were sampled between 28 and 32 weeks. An additional blood sample was obtained 4 to 6 months after delivery (baseline). The Mann-Whitney U test, the Friedman test, and the chi2 test were used. RESULTS: Patients in group 1 were characterized by increased factor VIIa levels, increased prothrombin fragment 1+2 levels, reduced factor XIIa levels, diminished functional urokinase-type plasminogen activator levels, and decreased levels of plasmin/alpha-2-plasmin inhibitor complexes. These abnormalities were more evident in patients in group 1 developing pre-eclampsia and/or intrauterine growth restriction. CONCLUSIONS: Impaired factor XIIa-dependent activation of fibrinolysis seems to be a key mechanism related to late-pregnancy complications in patients with the antiphospholipid syndrome.

Adult↗
PubMed2000

Human factor XII binding to the glycoprotein Ib-IX-V complex inhibits thrombin-induced platelet aggregation.

Factor XII deficiency has been postulated to be a risk factor for thrombosis suggesting that factor XII is an antithrombotic protein. The biochemical mechanism leading to this clinical observation is unknown. We have previously reported high molecular weight kininogen (HK) inhibition of thrombin-induced platelet aggregation by binding to the platelet glycoprotein (GP) Ib-IX-V complex. Although factor XII will bind to the intact platelet through GP Ibalpha (glycocalicin) without activation, we now report that factor XIIa (0. 37 microm), but not factor XII zymogen, is required for the inhibition of thrombin-induced platelet aggregation. Factor XIIa had no significant effect on SFLLRN-induced platelet aggregation. Moreover, an antibody to the thrombin site on protease-activated receptor-1 failed to block factor XII binding to platelets. Inhibition of thrombin-induced platelet aggregation was demonstrated with factor XIIa but not with factor XII zymogen or factor XIIf, indicating that the conformational exposure of the heavy chain following proteolytic activation is required for inhibition. However, inactivation of the catalytic activity of factor XIIa did not affect the inhibition of thrombin-induced platelet aggregation. Factor XII showed displacement of biotin-labeled HK (30 nm) binding to gel-filtered platelets and, at concentrations of 50 nm, was able to block 50% of the HK binding, suggesting involvement of the GP Ib complex. Antibodies to GP Ib and GP IX, which inhibited HK binding to platelets, did not block factor XII binding. However, using a biosensor, which monitors protein-protein interactions, both HK and factor XII bind to GP Ibalpha. Factor XII may serve to regulate thrombin binding to the GP Ib receptor by co-localizing with HK, to control the extent of platelet aggregation in vivo.

Amino Acid Sequence↗
PubMed1995

Activation of factors XII and VII induced in citrated plasma in the presence of contact surface.

Activated factor XII (XIIa), activated factor VII (VIIa) and factor VII coagulant activity (VIIc) were determined in non-treated and in treated (cold-incubated) citrated plasmas from women in late pregnancy and from norma volunteers. All three activities were higher in the non-treated plasmas from women in late pregnancy than from normal subjects. The incubation of citrated plasmas from women in late pregnancy, on ice for 24 hours, resulted in a many-fold increase of factor XIIa activity, factor VIIa levels and VIIc. The dilution of these plasmas resulted in a sharp decrease of all three activities in the post-incubation mixture, so that in the plasmas diluted 2:1 with buffer all three activities were similar to those in fresh plasmas. Similar incubations of diluted plasmas (1:1) from normal volunteers resulted in no increase of factor XIIa activity, factor VIIa levels and VIIc. However, the presence in the incubation mixture of micellar stearate resulted in a stearate concentration-dependent increase of all three activities in treated plasmas. Levels of factor XIIa activity and factor VIIa in the treated plasmas from both groups of subjects were highly correlated (r = 0.987; p < 0.001). There was also a highly significant correlation between VIIc and factor VIIa levels (0.989; p < 0.001). These results demonstrate that the in vitro increase in factor VIIa levels is due to the activation of the contact system of coagulation and is dependent on the potency of the contact surface. Moreover, VIIc over a wide range of values, observed in the present experiments, can provide an accurate measure of factor VIIa concentration.

Adult↗
PubMed1983

Mechanism of surface-mediated activation of bovine Factor XII and prekallikrein.

The mechanism of kaolin-mediated activation of bovine Factor XII was studied in the presence of prekallikrein and HMW kininogen. The activated enzymes were assayed using fluorogenic peptides, Boc-Glu (OBzl)-Gly-Arg-4-methylcoumaryl-7-amide (MCA) for Factor XIIa and Z-Phe-Arg-MCA for plasma kallikrein. The rates of activation of the zymogens were separately measured by blocking either of the active enzymes with specific inhibitors, corn inhibitor for Factor XIIa (Ki = 6.7 nM) and Trasylol for plasma kallikrein (Ki = 3.9 nM). The result was as follows: (1) At the early stage of the activation reaction, kallikrein activity was first generated after short lag time, and then Factor XIIa activity was generated with a sigmoidal curve. In the presence of corn inhibitor, the activation of prekallikrein was observed, but in the presence of Trasylol, the activation of Factor XII was not observed. In the presence of high concentration of Ala-Phe-Arg-Ch2Cl, which inactivates immediately both of the active enzymes, the cleavage of a single chain prekallikrein into the two chain form by Factor XII was shown by SDS-PAGE, using nonlabelled and tritiated prekallikrein. (2) The incubation of Factor XII alone in a quartz cuvette or in the presence of kaolin and HMW kininogen did not result in the activation of Factor XII. The concave upward curve due to an autocatalytic activation was not observed even after the addition of Factor XIIa to Factor XII preparation. Moreover, no structural change of Factor XII during the incubation with kaolin and HMW kininogen was shown by SDS-PAGE, using 3H-Factor XII. (3) The rates of activation of prekallikrein by Factor XII and by Factor XIIa were approximately the same at higher concentration of prekallikrein. However, at lower concentration of prekallikrein the rate of activation of prekallikrein by Factor XII was shown to be a sigmoidal curve and slower than that by Factor XIIa. These results indicate that the activation of bovine Factor XII is initiated by the attack of Factor XII on prekallikrein, followed by the reciprocal activation of Factor XII by plasma kallikrein generated. The autocatalytic activation of bovine Factor XII by Factor XIIa was not demonstrated.

Amino Acid Chloromethyl Ketones↗
PubMed1983

Sulfatide-dependent autoactivation of human blood coagulation Factor XII (Hageman Factor).

When purified human blood coagulation Factor XII (Hageman factor) is incubated with sulfatides at 37 degrees C, activation of Factor XII occurs as judged by the appearance of amidolytic activity towards the chromogenic substrate H-D-Pro-Phe-Arg-p-nitroanilide. Polyacrylamide gel electrophoresis studies using 125I-Factor XII as a marker show that the appearance of amidolytic activity correlates with Factor XII cleavage, that activation goes to completion and that virtually all Factor XIIa formed is present as the two-chain, 80,000 Mr form, alpha-Factor XIIa. Rigorous analysis of kinetic data establishes that, between 0.02 and greater than 90% of the reaction, the activation of Factor XII is described by a mechanism of autoactivation of Factor XII by Factor XIIa. The rate of autoactivation increases with increasing Factor XII concentrations at constant sulfatide levels but decreases with increasing sulfatide concentrations at constant levels of Factor XII. These findings suggest that the concentrations of Factor XII and Factor XIIa bound to the sulfatide surface determine the rate of autoactivation. Soybean trypsin inhibitor, Trasylol, and anti-prekallikrein antibodies have no influence on the rate of sulfatide-dependent autoactivation of Factor XII. Benzamidine inhibits autoactivation with an inhibitor constant, Ki, of 1.9 mM which is similar to the Ki of 1.5 mM for the enzyme, alpha-Factor XIIa. Thus, sulfatide-dependent activation of purified Factor XII is not due to contaminating proteases and is described by a second order mechanism of autoactivation due to the action of surface-bound Factor XIIa on surface-bound Factor XII.

Benzamidines↗
PubMed2003

Prevalence, causes, and characterization of factor XI inhibitors in patients with inherited factor XI deficiency.

Factor XI deficiency, an injury-related bleeding disorder, is rare worldwide but common in Jews in whom 2 mutations, Glu117Stop (type II) and Phe283Leu (type III), prevail. Mean factor XI activities in homozygotes for Glu117Stop and for Phe283Leu are 1 and 10 U/dL, respectively. Inhibitors to factor XI in patients with severe factor XI deficiency have been reported in a small number of instances. This study was undertaken to determine the prevalence of acquired inhibitors against factor XI in patients with severe factor XI deficiency, discern whether these inhibitors are related to specific mutations, and characterize their activity. Clinical information was obtained from unrelated patients with severe factor XI deficiency, and blood was analyzed for factor XI activity, inhibitor to factor XI, and causative mutations. Immunoglobulin G purified from patients with an inhibitory activity was tested for binding to factor XI, effects on activation of factor XI by factor XIIa and thrombin, and activation of factor IX by exogenous factor XIa. Of 118 Israeli patients, 7 had an inhibitor; all belonged to a subgroup of 21 homozygotes for Glu117Stop who had a history of plasma replacement therapy. Three additional patients with inhibitors from the United Kingdom and the United States also had this genotype and were exposed to plasma. The inhibitors affected factor XI activation by thrombin or factor XIIa, and activation of factor IX by factor XIa. The results imply that patients with a very low factor XI level are susceptible to development of an inhibitor following plasma replacement.

Aged↗
PubMed2004

Circulating cardiovascular risk factors in obstructive sleep apnoea: data from randomised controlled trials.

BACKGROUND: Obstructive sleep apnoea (OSA) is associated with high cardiovascular morbidity and mortality and is an independent risk factor for hypertension. Novel circulating cardiovascular risk markers enabling a more accurate prediction of cardiovascular risk have been identified. Examination of these markers may clarify the increased risk in OSA and contribute to an analysis of the benefits of treatment. METHODS: Plasma levels of total cholesterol and triglyceride and activated coagulation factors XIIa and VIIa, factors VII, VIII, XII, fibrinogen, thrombin-antithrombin (TAT), von Willebrand factor antigen (vWFAg), soluble P-selectin (sP-sel), and homocysteine were measured before and after treatment for 1 month with therapeutic or subtherapeutic (control) continuous positive airways pressure (CPAP) in 220 patients with OSA. RESULTS: Levels of activated coagulation factors XIIa, VIIa, TAT and sP-sel were higher in OSA patients at baseline than in unmatched controls, but did not fall with 1 month of therapeutic CPAP treatment. The raised sP-sel levels correlated only with body mass index (p = 0.002). There was a trend towards a significant fall in total cholesterol with therapeutic CPAP (p = 0.06) compared with the control group. In the therapeutic group there was a clinically significant mean fall in total cholesterol of 0.28 mmol/l (95% confidence interval 0.11 to 0.45, p = 0.001) which may reduce cardiovascular risk by about 15%. CONCLUSION: A number of activated coagulation factors are increased in untreated OSA patients, potentially contributing to vascular risk, but they do not fall with 1 month of CPAP treatment. Nasal CPAP may produce a clinically relevant fall in total cholesterol level, potentially reducing cardiovascular risk, but this needs to be verified in a larger prospective study.

Adult↗
PubMed1997

Partial identification of the Zn2+-binding sites in factor XII and its activation derivatives.

With the purpose of identifying the Zn2+-binding sites in factor XII, the effect of chemical modification of His, Glu and Asp residues, amino acids known to participate in the catalytic coordination binding of Zn2+ in a number of Zn2+-binding proteins, was analysed. The number of modifiable His residues in factor XII and alpha-factor XIIa was 16.0+/-0.7 and 17.3+/-0.7, respectively. When factor XII/alpha-factor XIIa was incubated with saturating concentrations of Zn2+ before the diethylpyrocarbonate modification of the His residues, these numbers were reduced to 6.3+/-0.1 and 8.21+/-0.5, indicating that ten and nine His residues, respectively, are involved in the binding. Analysis of the Zn2+-binding capacity of factor XII, alpha-factor XIIa and beta-factor XIIa showed that while factor XII contains four Zn2+-binding sites, alpha-factor XIIa had only three and beta-factor XIIa had none. Modification of the His residues resulted in a complete loss of Zn2+-binding while Asp/Glu modification resulted in loss of two and one Zn2+-binding sites in factor XII and alpha-factor XIIa, respectively. This suggests that two of the four sites in factor XII contain His residues, exclusively, while the two others are comprised of two His residues and one Asp/Glu residue. One of the latter is lost when factor XII is activated to alpha-factor XIIa. Two of the sites are suggested to be located at positions His40-His44 and His78-His82. The location of the remaining two sites are reduced to four possible positions.

Amino Acid Sequence↗
PubMed1988

Quantification of plasma factor XIIa-Cl(-)-inhibitor and kallikrein-Cl(-)-inhibitor complexes in sepsis.

Considerable evidence indicates that activation of the contact system of intrinsic coagulation plays a role in the pathogenesis of septic shock. To monitor contact activation in patients with sepsis, we developed highly sensitive radioimmunoassays (RIAs) for factor XIIa-Cl(-)-inhibitor (Cl(-)-Inh) and kallikrein-Cl(-)-Inh complexes using a monoclonal antibody (MoAb Kok 12) that binds to a neodeterminant exposed on both complexed and cleaved Cl(-)-Inh. Plasma samples were serially collected from 48 patients admitted to the intensive care unit because of severe sepsis. Forty percent of patients on at least one occasion had increased levels of plasma factor XIIa-Cl(-)-Inh (greater than 5 x 10(-4) U/mL) and kallikrein-Cl(-)-Inh (greater than 25 x 10(-4) U/mL), that correlated at a molar ratio of approximately 1:3. Levels of factor XII antigen in plasma and both the highest as well as the levels on admission of plasma factor XIIa-Cl(-)-Inh in 23 patients with septic shock were lower than in 25 normotensive patients (P = .015: factor XII on admission; P = .04: highest factor XIIa-Cl(-)-Inh; P = .01: factor XIIa-Cl(-)-Inh on admission). No significant differences in plasma kallikrein-Cl(-)-Inh or prekallikrein antigen were found between these patients' groups. Elevated Cl(-)-Inh complex levels were measured less frequently in serial samples from patients with septic shock than in those from patients without shock (P less than .0001). Based on these results, we conclude that plasma Cl(-)-Inh complex levels during sepsis may not properly reflect the extent of contact activation.

Adolescent↗