Some Data on Two Purified Kininogens From Human Plasma

Abstract

Work on the purification of the substrate in plasma for kinin-forming enzymes, kininogen, has been carried out in several laboratories. Habermann (1963) thus purified kininogen from bovine serum and Suzuki, Mizushima, Sato & Iwanaga (1965) purified a kininogen from bovine plasma. Habermann (1965Habermann ( , 1966 also studied the structure of the kinin-yielding polypeptides of his purified bovine kininogen (peptic kinin-yielding polypeptides). Pierce & Webster (1966) isolated two kininogens from human plasma. These kininogens were shown to be glycoproteins with molecular weights of about 50,000, as is also the case with the bovine kininogen (Habermann, 1963). Jacobsen (1966b) found that two different substrates for plasma kinin-forming enzymes could be separated from plasmas of several mammals. These substrates had different molecular dimensions and they differed in their reaction patterns towards various kininforming enzymes. The present paper reports the results of further purification with some characterization of these two substrates. The substrate with the larger molecular dimension has been designated substrate 1, and the other and smaller one has been designated substrate 2. The methods employed have been DEAE anion exchange chromatography, gel filtration, disc gel electrophoresis and sucrose gradient ultracentrifugation. METHODS Plasma. Citrated human plasma was obtained by using siliconized equipment throughout, as described by Jacobsen (1966b).Initial purification procedures for kininogens (anion exchange chromatography and gel filtration). The separation procedures previously described (Jacobsen, 1966a(Jacobsen, , 1966b have been modified and expanded, so as to include disc gel electrophoresis (see below). The initial procedure was a separation of fractions containing the two kininogens by chromatography of human citrated plasma on a DEAE anion exchanger. Each of the substrates was subsequently exposed to further purification procedures (see below and Table 1). For initial chromatography, plasma (25 ml.) was mixed with 0.05 M Tris-HCl buffer (pH 8) containing 0.25 M sodium chloride (12.5 ml.) and the mixture applied to a DEAE-Sephadex A 50 (A. B. Pharmacia, Uppsala, Sweden) column (18 x 1.8 cm) equilibrated with 0.05 M Tris-HCl buffer (pH 8) containing 0.2 M sodium chloride. Substrate 1 is retained on the column while substrate 2 is eluted with the equilibrating buffer. Substrate 1 could then be eluted with a 0.05 M Tris-HCl buffer (pH 8) containing 0.35 M sodium chloride. The effluent containing substrate 2 was treated with " massive contact exposure " in order to eliminate the plasma kallikrein activity (Jacobsen, 1966b), and then applied to a DEAE-Sephadex A 50 column (10 x 1 cm) equilibrated with 0.05 M Tris-HCI buffer (pH 8) containing 0.1 M sodium chloride. Substrate 2 was eluted from this column with a buffer containing 0.25 M sodium chloride. Buffer concentrations in the samples were always adjusted to that of the buffer used for equilibration in the following purification procedure. S. JACOB...

Williams trait. Human kininogen deficiency with diminished levels of plasminogen proactivator and prekallikrein associated with abnormalities of the Hageman factor-dependent pathways.

“…Although it is clear from the studies of Spragg and Austen that the majority of plasma kininogen is of low molecular weight (49), this does not appear to be the sole form of native human kininogen. Previous authors have described a human high molecular weight kininogen that possesses different kinetic properties when activated by kallikrein (15,20,50,51). It is clear from our studies that such a high molecular weight kininogen can be separated from low molecular weight kininogen and this kininogen has a special function in the early steps of the Hageman factor-dependent pathways.…”

Flaujeac factor deficiency. Reconstitution with highly purified bovine high molecular weight-kininogen and delineation of a new permeability-enhancing peptide released by plasma kallikrein from bovine high molecular weight-kininogen.

“…The existence of kininogens of high molecular weight was first described by Jacobsen (14) and Jacobsen and Kriz (15). In contrast to LMW-kininogen, the HMW-kininogen showed a preferential susceptibility to plasma kallikrein.…”

Function and Immunochemistry of Prekallikrein-High Molecular Weight Kininogen Complex in Plasma

“…Close functional relationships, however, do exist between these two proteins. The activated form of prekallikrein, kallikrein, is the enzyme which releases bradykinin from HMW kininogen (6), whereas HMW kininogen in turn potentiates the activation of prekallikrein on a surface by Factor XII (7,8) or in the fluid phase by Factor XII fragments (XIIf) (9). The former activity may involve the increased binding ofprekallikrein to the surface when complexed with HMW kininogen.…”