Sobel J H, Trakht I, Wu H Q, Rudchenko S, Egbring R
Department of Medicine, College of Physicians & Surgeons of Columbia University, New York, NY 10032, USA.
Blood. 1995 Aug 1;86(3):989-1000.
The fibrinogen structural variant, Marburg (A alpha 1-460B beta gamma)2, is comprised of normal B beta and gamma chains but contains severely truncated A alpha chains that are missing approximately one half of their factor XIIIa cross-linking domain. Immunochemical studies of fibrin(ogen) Marburg were conducted to characterize the degree to which deletion of a defined A alpha-chain segment, A alpha 461-610, can affect the process of fibrin stabilization, ie, the factor XIIIa-mediated covalent interaction that occurs between alpha chains of neighboring fibrin molecules and between alpha chains and alpha 2 antiplasmin (alpha 2PI). The ability of Marburg (and control) alpha chains to serve as a substrate for factor XIIIa and undergo cross-linking was examined in an in vitro plasma clotting system. The capacity for alpha-chain cross-linking was evaluated both as the covalent incorporation of the small synthetic peptide, NQEQVSPLTLLK (which represents the first 12 amino acids of alpha 2PI and includes the factor XIIIa-sensitive glutamine residue responsible for the cross-linking of alpha 2PI to fibrin), and as the appearance of native (ie, natural), high-molecular-weight, cross-linked alpha-chain species. Antibodies specific for the (A)alpha and gamma/gamma-gamma chains of fibrin(ogen) and for the peptide and its parent protein, alpha 2PI (68 kD), were used as immunoblotting probes to visualize the various cross-linked products formed during in vitro clotting. Recalcification of Marburg plasma in the presence of increasing concentrations of peptide resulted in the formation of peptide-decorated Marburg alpha-chain monomers. Their size at the highest peptide concentration examined indicated the incorporation of a maximum of 3 to 4 mol of peptide per mole of alpha-chain. In the absence of alpha 2PI 1-12 peptide, the alpha chains of Marburg fibrin cross-linked to form oligomers and polymers, as well as heterodimers that included alpha 2PI. Both the peptide-decorated monomers and the native cross-linked alpha-chain species of Marburg fibrin were smaller than their control plasma counterparts, consistent with the truncated structure of the parent Marburg A alpha chain. Collectively, the findings indicate that, although deletion of the A alpha chain region no. 461-610 in fibrinogen Marburg prevents formation of an extensive alpha polymer network (presumably due to the absence of critical COOH-terminal lysine residues), it does not interfere with initial events in the fibrin stabilization process, namely, factor XIII binding and the ability of alpha chains to undergo limited cross-linking to one another and to alpha 2PI.
纤维蛋白原结构变体马尔堡型（Aα1 - 460Bβγ）2由正常的Bβ链和γ链组成，但含有严重截短的Aα链，其缺失了大约一半的因子XIIIa交联结构域。对纤维蛋白原马尔堡型进行了免疫化学研究，以确定特定Aα链片段Aα461 - 610的缺失在多大程度上会影响纤维蛋白稳定过程，即因子XIIIa介导的相邻纤维蛋白分子α链之间以及α链与α2抗纤溶酶（α2PI）之间发生的共价相互作用。在体外血浆凝血系统中检测了马尔堡型（及对照）α链作为因子XIIIa底物并进行交联的能力。通过小合成肽NQEQVSPLTLLK（代表α2PI的前12个氨基酸，包括负责α2PI与纤维蛋白交联的因子XIIIa敏感谷氨酰胺残基）的共价掺入以及天然（即正常）高分子量交联α链物种的出现来评估α链交联能力。针对纤维蛋白原的（A）α链和γ/γ - γ链以及该肽及其亲本蛋白α2PI（68kD）的特异性抗体用作免疫印迹探针，以可视化体外凝血过程中形成的各种交联产物。在存在浓度不断增加的肽的情况下对马尔堡型血浆进行再钙化，导致形成肽修饰的马尔堡型α链单体。在检测的最高肽浓度下其大小表明每摩尔α链最多掺入3至4摩尔肽。在没有α2PI 1 - 12肽的情况下，马尔堡型纤维蛋白的α链交联形成寡聚体、聚合物以及包含α2PI的异二聚体。马尔堡型纤维蛋白的肽修饰单体和天然交联α链物种均小于其对照血浆对应物，这与亲本马尔堡型Aα链的截短结构一致。总体而言，研究结果表明，尽管纤维蛋白原马尔堡型中Aα链区域461 - 610的缺失阻止了广泛的α聚合物网络的形成（可能是由于关键的COOH末端赖氨酸残基缺失），但它并不干扰纤维蛋白稳定过程中的初始事件即因子XIII结合以及α链彼此之间以及与α2PI进行有限交联的能力。
