title: Purification of high molecular weight kininogen and the role of this agent in blood coagulation.
abstract: Recent studies of individuals with high molecular weight (HMW) kininogen deficiency established the importance of this plasma protein for in vitro initiation of blood coagulation. In the present study, HMW-kininogen was highly purified from human plasma by monitoring its clot-promoting activity, using Fitzgerald trait plasma as a substrate. This preparation of HMW-kininogen revealed a single band on sodium dodecyl sulfate-polyacrylamide gel electrophoresis (mol wt: 120,000) and released 1% of its weight as bradykinin upon incubation with plasma kallikrein. HMW-kininogen specifically repaired impaired surface-mediated plasma reactions of Fitzgerald trait plasma, but did not affect those of Hageman trait and Fletcher trait plasma. Kinin release from HMW-kininogen by trypsin, but not by plasma kallikrein, resulted in total loss of clot-promoting activity. No inhibitors of coagulation were found when all kinin activity was removed from HMW-kininogen by trypsin. The roles of HMW-kininogen, Hageman factor (HF, Factor XII), plasma prekallikrein (Fletcher factor), and plasma thromboplastin antecedent (PTA, Factor XI) in blood coagulation were studied in a purified system. HMW-kininogen was absolutely required for activation of PTA by HF and ellagic acid. The yield of activated PTA was proportional to the amount of HF, HMW-kininogen, and PTA in the mixtures, suggesting that, to activate PTA, these three proteins might form a complex in the presence of ellagic acid. No fragmentation of HF was found under these conditions. In contrast to HF, HF-fragments (mol wt: 30,000) activated PTA in the absence of HMW-kininogen and ellagic acid. Thus, it appears that in the present study PTA was activated in two distinct ways. Which pathway is the major one in whole plasma remains to be determined.
mesh_terms: D001777:Blood Coagulation / Q000187:drug effects; D004610:Ellagic Acid / Q000494:pharmacology; D005172:Factor XI / Q000378:metabolism; D005174:Factor XII / Q000494:pharmacology; D006801:Humans; D066298:In Vitro Techniques; D007610:Kallikreins / Q000494:pharmacology; D007704:Kininogens / Q000302:isolation & purification / Q000502:physiology*; D008722:Methods; D014357:Trypsin / Q000494:pharmacology
chemical_list: D007704:Kininogens; D004610:Ellagic Acid; D005174:Factor XII; D005172:Factor XI; D007610:Kallikreins; D014357:Trypsin