Váradi A, Scheraga H A
Biochemistry. 1986 Feb 11;25(3):519-28. doi: 10.1021/bi00351a001.
We have isolated an intermediate plasmic degradation product, D2, of fibrinogen that does not inhibit the polymerization of fibrin monomer but does bind Ca2+. Fibrinogen was digested to a limited extent with plasmin in the presence of Ca2+, and a "large" fragment D (fragment D1A) was isolated with a gamma-chain remnant consisting of residues 63-411. Fragment D1A was digested further in the presence of Ca2+, yielding fragment D1 (with its gamma-chain containing residues 86-411). The digestion of fragment D1 [in the presence of ethylene glycol bis(beta-aminoethyl ether)-N,N,N',N'-tetraacetic acid (EGTA) to complex Ca2+] led to a gradual shortening of the carboxyl-terminal portion of the gamma-chain. Fragment D2 (with its gamma-chain containing residues 86-335/356) was isolated from an intermediate digest in the presence of EGTA. The Lys-338-Cys-339 peptide bond of the gamma-chain is intact in this preparation of D2, even though it is split in the isolated peptide gamma303-355 (with an intact disulfide bond at Cys-326-Cys-339). Fragment D2 does not interfere with the polymerization of fibrin monomer, whereas fragment D1 is a potent inhibitor of this polymerization. We conclude that the gamma-chain segment 356/357-411, present in fragment D1 but absent from fragment D2, is essential for maintenance of a polymerization site located in the outer (D) nodule of fibrinogen. This segment (356/357-411) is longer than two shorter ones reported earlier [Olexa, S.A., & Budzynski, A. Z. (1981) J. Biol. Chem. 256, 3544-3549; Horwitz, B.H., Váradi, A., & Scheraga, H.A. (1984) Proc. Natl. Acad. Sci. U.S.A. 81, 5980-5984]; the data for the earlier reports are reinterpreted here. Finally, fragment D2 possesses a single Ca2+ binding site, as revealed by equilibrium dialysis binding studies. Since fragment D3 (with its gamma-chain containing residues 86-302) fails to bind Ca2+, we conclude that segment gamma 303-355/356 plays a crucial role in Ca2+ binding.
我们分离出了纤维蛋白原的一种中间血浆降解产物D2，它不抑制纤维蛋白单体的聚合，但能结合Ca2+。在Ca2+存在的情况下，用纤溶酶对纤维蛋白原进行有限程度的消化，分离出一个“大”的D片段（片段D1A），其γ链残余部分由63 - 411位残基组成。在Ca2+存在的情况下，对片段D1A进一步消化，产生片段D1（其γ链包含86 - 411位残基）。在乙二醇双（β - 氨基乙基醚）- N,N,N',N'-四乙酸（EGTA）存在下对片段D1进行消化（以络合Ca2+），导致γ链羧基末端部分逐渐缩短。在EGTA存在的情况下，从中间消化产物中分离出片段D2（其γ链包含86 - 335/356位残基）。在D2的这种制备中，γ链的Lys - 338 - Cys - 339肽键是完整的，尽管它在分离的肽γ303 - 355中被切断（在Cys - 326 - Cys - 339处有完整的二硫键）。片段D2不干扰纤维蛋白单体的聚合，而片段D1是这种聚合的有效抑制剂。我们得出结论，片段D1中存在但片段D2中不存在的γ链片段356/357 - 411对于维持位于纤维蛋白原外部（D）结节中的聚合位点至关重要。该片段（356/357 - 411）比之前报道的两个较短片段长[Olexa, S.A., & Budzynski, A. Z. (1981) J. Biol. Chem. 256, 3544 - 3549; Horwitz, B.H., Váradi, A., & Scheraga, H.A. (1984) Proc. Natl. Acad. Sci. U.S.A. 81, 5980 - 5984]；这里对早期报道的数据进行了重新解释。最后，平衡透析结合研究表明，片段D2具有单个Ca2+结合位点。由于片段D3（其γ链包含86 - 302位残基）不能结合Ca2+，我们得出结论，γ链片段303 - 355/356在Ca2+结合中起关键作用。
Zotero 插件