Synthesis of truncated amino-terminal trimers of thrombospondin.
Synthesis of truncated amino-terminal trimers of thrombospondin.
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DOI：
10.1021/bi00240a028
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发表时间：
1991-07
期刊：
影响因子：
2.9
通讯作者：
J. Sottile;J. Selegue;D. Mosher
中科院分区：
文献类型：
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作者：
J. Sottile;J. Selegue;D. Mosher
血小板反应蛋白（TSP）是一种450 kda的糖蛋白，由三个相同的二硫键结合亚基（1152个氨基酸）组成，它们在肝素结合氨基末端的球状结构域附近结合在一起。残基277 （TSP-277）或381 （TSP-381）截断的TSP在COS细胞或昆虫细胞中表达时，主要由二硫键结合三聚体组成。此外，TSP-381与内源性COS细胞TSP形成异源三聚体。对TSP和残基220和237之间的蛋白水解敏感区域的截短突变体进行切割，得到单体氨基末端片段。Cys-252和Cys-256是残基238和277之间唯一的半胱氨酸，因此必须在亚基之间架起桥梁。将Cys-252和Cys-256转化为甘氨酸的TSP-381能被COS细胞有效分泌，但只有一小部分蛋白以二硫键结合三聚体的形式存在。预测残基258和283之间的TSP序列形成一个两族α -螺旋。我们认为，TSP三聚体的组装涉及形成一种螺旋状线圈结构，这种结构通过形成二硫化物来稳定。
Thrombospondin (TSP) is a 450-kDa glycoprotein that is comprised of three identical disulfide-bonded subunits (1152 amino acids) held together near the heparin-binding amino-terminal globular domains. TSP truncated at residue 277 (TSP-277) or 381 (TSP-381) consisted largely of disulfide-bonded trimers when expressed in COS cells or insect cells. In addition, TSP-381 formed heterotrimers with endogenous COS cell TSP. Cleavage of TSP and the truncated mutants in the proteolytically sensitive region between residues 220 and 237 yielded monomeric amino-terminal fragments. Cys-252 and Cys-256 are the only cysteines between residues 238 and 277 and therefore must bridge among subunits. TSP-381 in which Cys-252 and Cys-256 were changed to glycine was secreted efficiently by COS cells but with only a minor portion of the protein in the form of disulfide-bonded trimers. The sequence of TSP between residues 258 and 283 is predicted to form an amphiphatic alpha-helix. We suggest that assembly of TSP trimers involves formation of an alpha-helical coiled-coil structure which is stabilized by formation of disulfides.