Luczak J A, Redick S D, Schwarzbauer J E
Department of Molecular Biology, Princeton University, New Jersey 08544-1014, USA.
J Biol Chem. 1998 Jan 23;273(4):2073-7. doi: 10.1074/jbc.273.4.2073.
Tenascin-C is a large, multimeric extracellular matrix protein that is found in a variety of tissues and can have profound effects on cell adhesion. It is secreted from cells as a hexamer of six identical chains called a hexabrachion. Disulfide bonding among tenascin subunits mediates intracellular assembly into hexamers. The amino-terminal assembly domain consists of heptad repeats and at least six cysteine residues (Cys-64, -111, -113, -140, -146, -147) that could be involved in multimerization. We have now determined the requirements for these cysteine residues during hexamer assembly. Our results show that only Cys-64 is required to form the hexameric structure. Mutation of Cys-64 to glycine resulted in release of trimer intermediates, which probably form via the heptad repeats, but no hexamers were secreted. In contrast, individual or pairs of mutations of each of the other cysteines had no effect on tenascin hexamer formation, and inclusion of any other cysteine mutations along with C64G did not further disrupt the multimer pattern. However, when all six cysteines were mutated, monomers were the major extracellular form. Together, these results show that trimers are an intermediate of tenascin-C assembly and that Cys-64 is essential for formation of hexabrachions.
腱生蛋白-C是一种大型多聚体细胞外基质蛋白，存在于多种组织中，对细胞黏附可产生深远影响。它作为由六条相同链组成的六聚体（称为六臂体）从细胞中分泌出来。腱生蛋白亚基之间的二硫键介导细胞内组装成六聚体。氨基末端组装结构域由七肽重复序列和至少六个半胱氨酸残基（Cys-64、-111、-113、-140、-146、-147）组成，这些残基可能参与多聚化。我们现在已经确定了六聚体组装过程中这些半胱氨酸残基的需求。我们的结果表明，仅需Cys-64来形成六聚体结构。将Cys-64突变为甘氨酸导致三聚体中间体的释放，三聚体可能通过七肽重复序列形成，但没有六聚体被分泌。相比之下，其他每个半胱氨酸的单个或成对突变对腱生蛋白六聚体的形成没有影响，并且将任何其他半胱氨酸突变与C64G一起引入也不会进一步破坏多聚体模式。然而，当所有六个半胱氨酸都发生突变时，单体成为主要的细胞外形式。总之，这些结果表明三聚体是腱生蛋白-C组装的中间体，并且Cys-64对于六臂体的形成至关重要。
