Abstract. The three unique polypeptide chains of human fibrinogen differ significantly in molecular weight. Cross-linkage of fibrin by fibrin-stabilizing factor results in the rapid formation of cross-links between y-chains and a slower formation of cross-links between a-chains. p-Chains are not involved directly in the cross-linking of fibrin. Reduced, cross-linked fibrin contains uncrosslinked i-chains, dimers of y-chain, and higher polymers of a-chain. Although it is uncertain whether the y-,y dimers are formed by chains in different molecules of fibrin, the polymers of a-chain in fibrin can only be accounted for by crosslinkage of a-chains in different molecules. The nature of cross-linkage among the subunits in fibrin can account well for the three-dimensional, covalent structure of cross-linked, insoluble fibrin.Human fibrinogen, which has a molecular weight of approximately 330,000, contains three pairs of different polypeptide subunits (a, Al, y) and may be designated by the polypeptide chain formula a38272.1-4 The three chains are combined covalently through disulfide bonds and differ in amino acid sequence and molecular weight. It has been estimated that the molecular weights of the a-, (3-, and 7-chains are 63,500, 56,000, and 47,000, respectively.4 Fibrin is formed when thrombin acts on fibrinogen and cleaves specific peptide bonds in the a-and ,8-chains, thereby releasing low molecular weight peptides. Thrombin action alone produces fibrin which is less soluble than fibrinogen but is soluble at low pH or in concentrated solutions of urea or guanidine hydrochloride. Such fibrin is termed noncross-linked, soluble fibrin. Soluble fibrin is converted to cross-linked, insoluble fibrin by a transpeptidase, termed fibrin-stabilizing factor (FSF), which catalyzes the formation of new amide bonds between e-amino groups of lysine and carboxamido groups of glutamine in the subunit chains.5 Thus, cross-linked, insoluble fibrin, after reduction of its disulfide bonds, contains higher molecular weight subunits than fibrinogen. Chen and Doolittle6 concluded that reduced, cross-linked bovine fibrin contains y-7y as well as a--y dimers, whereas Lorand and co-workers found mainly a--y dimers.7 8 Similarly, Takagi and Iwanaga9 suggest that two to four 7-chains are cross-linked in insoluble fibrin.We wish to report here the results of studies on the subunit structure of human fibrinogen and cross-linked, insoluble, human fibrin. During the course of the 738

No abstract

1. An enzyme that catalyses the transfer of sulphate from adenosine 3'-phosphate 5'[(35)S]-sulphatophosphate to l-tyrosine methyl ester and tyramine was purified approx. 70-fold from female rat livers. 2. The partially purified preparation is still contaminated with adenosine 3'-phosphate 5'-sulphatophosphate-phenol sulphotransferase (EC 2.8.2.1), but a partial separation of the two enzymes can be achieved by chromatography on columns of Sephadex G-200 and DEAE-Sephadex. 3. The enzyme responsible for the sulphation of l-tyrosine methyl ester and tyramine is activated by dithiothreitol, 2-mercaptoethanol and GSH, the degree of activation being more marked with preparations previously stored at 0 or -10 degrees C. In contrast, the enzymic sulphation of p-nitrophenol is inhibited by all three thiols. Again, there is a quantitative difference in the degree of inhibition of the two enzymes by o-iodosobenzoate, p-chloromercuribenzoate, N-ethylmaleimide and iodoacetate. 4. Mixed-substrate experiments support the hypothesis that the enzyme responsible for the sulphation of l-tyrosine methyl ester and tyramine is separate from that responsible for the sulphation of p-nitrophenol. However, p-nitrophenol is a potent inhibitor of the sulphation of both tyrosyl derivatives whereas these latter compounds have no effect on the sulphation of p-nitrophenol.

No abstract

No abstract