An amino acid polymorphism in histidine-rich glycoprotein (HRG) explains 59% of the variance in plasma HRG levels

A pedigree-based maximum likelihood method developed by Lange et al. (12) was used to study the contribution of a newly defined di-allelic polymorphism in histidine-rich glycoprotein (HRG) to the plasma levels of HRG. In four families (n = 99) and 20 volunteers we found a heritability of 70%, an age effect of 3% and an effect of individual environmental factors of 27%. These results are remarkably similar to the results found in a previous parent-twin study in which a heritability of 69% and an effect of random environment of 31% was found. The overall genetic influence in the present study can be subdivided into an effect of 59% by the HRG phenotype and 11% by residual genetic factors. The influence of the HRG phenotype of 59% can entirely be explained by adding up the effect of the two alleles that make up the phenotype. These results indicate a codominant inheritance pattern of HRG levels in which the genetic influence can almost completely be ascribed to the additive effect of the di-allelic HRG locus whereas only a small part is due to other loci

Fibrinolytic properties of activated factor XII

Activated factor XII (FXIIa), the initiator of the contact activation system, has been shown to activate plasminogen in a purified system. However, the quantitative role of FXIIa as a plasminogen activator in (factor XH-dependent) fibrinolysis in plasma is still unclear. The aim of our study was to investigate plasminogen activator activity (PAA) of both FXIIa in a purified system and endogenous FXIIa in plasma by measuring fibrinolysis in a clot lysis assay. During activation of purified FXII by trypsin, PAA was generated and remained stable even when FXIIa was completely cleaved to smaller fragments. Far more PAA was generated during autoactivation of purified FXII in the presence of dextran sulphate (DXS), a negatively charged component. However, PAA of autoactivated FXIIa decreased when it was cleaved to smaller fragments and reached a plateau comparable to PAA after activation by trypsin, while amidolytic activity of FXIIa remained constant. The same result was found when purified FXII was activated by kallikrein in the presence of DXS, while PAAwas reduced even more rapidly due to faster cleavage of FXII. This finding suggested that DXS may potentiate PAA of FXIIa in the clot lysis assay and not of the FXIIa fragments, missing the binding domain for DXS. Indeed, DXS was found to potentiate PAA of FXIIa more than tenfold according to a template model. Factor XII-dependent fibrinolysis is routinely measured in a dextran sulphate euglobulin fraction (DEF) of plasma and accounts for about 50% of total PAA. Since DXS was found to potentiate PAA of purified FXIIIa, FXI1 itself may be an important candidate for factor XIIdependent PAA in the DEF, a milieu in which DXS is present. Therefore, a DEF was prepared from normal human plasma and immunodepleted of FXIIa using an anti-FXII IgG Sepharose column. About 20% of factor XII-dependent PAA in the DEF could be ascribed to FXIIa. This study demonstrates that the fibrinolytic activity of FXIIa is substantially potentiated by DXS. Due to this potentiation, FXIIa significantly contributes to factor XII-dependent fibrinolysis in plasma as measured in a dextran sulphate euglobulin fraction

Escherichia coli Phosphoenolpyruvate Dependent Phosphotransferase System. Complete Purification of Enzyme I by Hydrophobic Interaction Chromatography

We have observed that EI possesses extremely hydrophobic surface regions. In light of this property, a high yield, rapid procedure has been developed for the complete purification of EI using solely hydrophobic interaction chromatography on commercially available resins. The entire procedure can be completed in 4-5 days with a 55-65% recovery of EI activity and an 870-fold purification.

The Escherichia coli Phosphoenolpyruvate-Dependent Phosphotransferase System: Observation of Heterogeneity in the Amino Acid Composition of HPr

Resonances of the aromatic protons of tyrosine have been observed in the proton nuclear magnetic resonance (1H NMR) spectrum of purified HPr from Escherichia coli. Analysis of the NMR spectrum of native HPr suggests that the tyrosine is located in a single position in the secondary structure and that this position is on the interior of the molecule inaccessible to solvent. Previous reports suggested that E. coli HPr contained no tyrosine. In contrast, we find, by amino acid analysis and ultraviolet and NMR spectroscopy, that E. coli HPr does contain tyrosine but at a subintegral level of 0.5 ± 0.1 mol of tyrosine per mol of HPr.

Escherichia coli Phosphoenolpyruvate Dependent Phosphotransferase System. NMR Studies of the Conformation of HPr and P-HPr and the Mechanism of Energy Coupling

1H and 31P nuclear magnetic resonance investigations of the phosphoprotein intermediate P-HPr and the parent molecule HPr of the E. coli phosphoenolpyruvate dependent phosphotransferase system (PTS) show that HPr can exist in two conformations. These conformations influence the protonation state of the reactive histidine residue, thereby determining the reaction pathway in the phosphoryl group transfer step. A general mechanism is proposed for the energy-coupling process in the PTS.