A Convenient Large-Scale Preparation of High Molecular Weight Kininogen from Human Plasm

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Abstract
Human high molecular weight (HMW) kininogen was purified by chromatography on DEAE-Sephadex A-50 and carboxymethyl-Sephadex C-50, followed by gel filtration on Sephadex G-50. From 5 l fresh human plasma .apprx. 120 mg HMW kininogen was obtained. The yield was 40%. The preparation had a specific activity of 14 .mu.g bradykinin equivalent/absorbance at 280 nm of ol. Upon polyacrylamide disc gel electrophoresis HMW kininogen was separated into 2 close bands, whereas only 1 band with an apparent MW of 120,000 was obtained in sodium dodecyl sulfate electrophoresis. Both protein fractions separated in disc gel electrophoresis released kinins upon incubation with kallikreins. The purified HMW kininogen had an isoelectric point of 4.65 when measured by isoelectric focusing. The amino acid composition of the purified HMW kininogen is given. The amino terminus of the molecule is blocked. In oligomerization studies adducts with MW up to 810,000 were obtained. HMW kininogen gave a single precipitin are in immunoelectrophoresis with antiserum directed against HMW kininogen.

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