Analysis of functional domains within adhesive proteins with the aim of controlling the interaction between cells and collagens

胶原蛋白是细胞外基质中最常见的蛋白质之一。在细胞外基质中，有像胶原这样的蛋白质以细胞外基质蛋白的形式存在，并且有与这些细胞外基质蛋白相关的蛋白质，并在结合状态下发挥各自的功能。我们一直对后一类蛋白质感兴趣，并一直在努力定位这些蛋白质中的功能结构域。以下是我们在过去三年中利用这项研究拨款进行的研究的主要发现。(1)我们从与I型胶原结合的血小板中分离出一种蛋白质。通过N端氨基酸序列分析，我们将该蛋白鉴定为von Willebrand因子前多肽(pp-vWF)，被称为von Willebrand抗原II。我们成功地将胶原结合域缩小到843个氨基酸残基中的10个残基部分Trp620-Ser629。作为成熟的von Willebrand因子，由c-…衍生而来。更常见的pp-vWF前体也与I型胶原结合，我们对这两种蛋白与胶原结合的特性进行了比较。我们得出的结论是，它们识别胶原分子中不同的结合部位。我们发现pp-vWF不仅存在于α颗粒中，而且存在于血小板表面。这可能表明它在血小板与内皮下胶原蛋白的黏附过程中起作用。我们发现pp-vWF通过凝血因子XIIIA或组织转谷氨酰胺酶与细胞外基质蛋白层粘连蛋白交联。然后我们研究了另一种胶原结合蛋白-凝血酶敏感蛋白(TSP)。TSP与pp-vWF的不同之处在于，它优先与天然的V型胶原结合，而不是与pp-vWF不同的I型胶原结合。由于我们有一个未来的计划来比较这两个胶原结合域，我们研究了TSP的V型胶原结合域。我们能够将该结构域定位到450 kDa分子中由80个氨基酸残基组成的部分。较少

Collagen is one of the most common proteins found in extra-cellular matrix (ECM). In ECM there are proteins like collagen which are inherently present there as ECM proteins, and there are proteins which are associated with these ECM proteins and exert respective functions in the associated state. We have been interested in the latter kind of proteins and have been trying to locate the functional domains within these proteins. Followings are the main findings we have made from the research carried out during the last three years using this research grant.(1). We isolated a protein from platelets bound to type I collagen. We identified this protein by N-terminal amino acid sequence analyses as propolypeptide of von Willebrand factor (pp-vWF) which had been called von Willebrand antigen II.(2). We succeeded in narrowing down the collagen binding domain to a ten-residue portion Trp620-Ser629 out of 843 amino acid residues.(3). As the mature von Willebrand factor which is derived from the c … More ommon precursor for pp-vWF is also known to bind to type I collagen, we made a comparison between these two proteins in terms of binding characteristics to the collagen. We came to a conclusion that they recognize different sites in the collagen molecule for the binding.(4). We found that pp-vWF was present not only in alpha-granules but also on the surface of platelets. This may indicate that it plays a role during the adhesion of platelets to collagen present in subendothelium.(5). We found pp-vWF cross-linked to an ECM protein laminin via blood coagulation factor XIIIa or tissue transglutaminase.(6). We then investigated another collagen-binding protein thrombospondin (TSP). TSP was different from pp-vWF in that it preferentially bound to native type V collagen, rather than type I collagen unlike pp-vWF.As we have a future plan of making a comparison between these two collagen binding domains, we studied the type V collagen-binding domain of TSP.We were able to locate the domain to a portion comprised of 80 amino acid residues out of the 450-kDa molecule. Less