Litvinov Rustem I, Yakovlev Sergiy, Tsurupa Galina, Gorkun Oleg V, Medved Leonid, Weisel John W
Department of Cell and Developmental Biology, University of Pennsylvania School of Medicine, Philadelphia, Pennsylvania 19104-6058, USA.
Biochemistry. 2007 Aug 7;46(31):9133-42. doi: 10.1021/bi700944j. Epub 2007 Jul 13.
The carboxyl-terminal regions of the fibrinogen Aalpha chains (alphaC regions) form compact alphaC-domains tethered to the bulk of the molecule with flexible alphaC-connectors. It was hypothesized that in fibrinogen two alphaC-domains interact intramolecularly with each other and with the central E region preferentially through its N-termini of Bbeta chains and that removal of fibrinopeptides A and B upon fibrin assembly results in dissociation of the alphaC regions and their switch to intermolecular interactions. To test this hypothesis, we studied the interactions of the recombinant alphaC region (Aalpha221-610 fragment) and its subfragments, alphaC-connector (Aalpha221-391) and alphaC-domain (Aalpha392-610), between each other and with the recombinant (Bbeta1-66)2 and (beta15-66)2 fragments and NDSK corresponding to the fibrin(ogen) central E region, using laser tweezers-based force spectroscopy. The alphaC-domain, but not the alphaC-connector, bound to NDSK, which contains fibrinopeptides A and B, and less frequently to desA-NDSK and (Bbeta1-66)2 containing only fibrinopeptides B; it was poorly reactive with desAB-NDSK and (beta15-66)2 both lacking fibrinopeptide B. The interactions of the alphaC-domains with each other and with the alphaC-connector were also observed, although they were weaker and heterogeneous in strength. These results provide the first direct evidence for the interaction between the alphaC-domains and the central E region through fibrinopeptide B, in agreement with the hypothesis given above, and indicate that fibrinopeptide A is also involved. They also confirm the hypothesized homomeric interactions between the alphaC-domains and display their interaction with the alphaC-connectors, which may contribute to covalent cross-linking of alpha polymers in fibrin.
纤维蛋白原Aα链的羧基末端区域（αC区域）形成紧密的αC结构域，通过柔性的αC连接子与分子主体相连。据推测，在纤维蛋白原中，两个αC结构域通过Bβ链的N末端优先在分子内相互作用，并与中央E区域相互作用，并且在纤维蛋白组装时纤维蛋白肽A和B的去除导致αC区域解离并转变为分子间相互作用。为了验证这一假设，我们使用基于激光镊子的力谱技术，研究了重组αC区域（Aα221 - 610片段）及其亚片段αC连接子（Aα221 - 391）和αC结构域（Aα392 - 610）之间以及它们与重组（Bβ1 - 66）2和（β15 - 66）2片段以及对应于纤维蛋白（原）中央E区域的NDSK之间的相互作用。αC结构域而非αC连接子与含有纤维蛋白肽A和B的NDSK结合，与仅含有纤维蛋白肽B的desA - NDSK和（Bβ1 - 66）2结合的频率较低；它与既缺乏纤维蛋白肽B的desAB - NDSK和（β15 - 66）2反应性较差。还观察到αC结构域之间以及它们与αC连接子之间的相互作用，尽管它们较弱且强度不均一。这些结果为αC结构域与中央E区域通过纤维蛋白肽B相互作用提供了首个直接证据，与上述假设一致，并表明纤维蛋白肽A也参与其中。它们还证实了αC结构域之间假设的同聚体相互作用，并展示了它们与αC连接子的相互作用，这可能有助于纤维蛋白中α聚合物的共价交联。
