Griffin P R, Kumar S, Shabanowitz J, Charbonneau H, Namkung P C, Walsh K A, Hunt D F, Petra P H
Department of Chemistry, University of Virginia, Charlottesville 22901.
J Biol Chem. 1989 Nov 15;264(32):19066-75.
The amino acid sequence of the sex steroid-binding protein (SBP or SHBG) of rabbit serum, specific for binding testosterone and 5 alpha-dihydrotestosterone, was determined using a complementary combination of mass spectrometric and Edman degradation techniques. The monomeric unit of the homodimeric protein is a single chain glycopeptide of 367 amino acid residues, with N-linked oligosaccharide side chains at Asn-345 and Asn-361 and disulfide bonds connecting Cys-158 to Cys-182 and Cys-327 to Cys-355. The polypeptide molecular weight of the monomer calculated from the sequence is 39,769. The molecular weight of the homodimer including 9% carbohydrate is 87,404. The sequence contains a relatively hydrophobic segment between Trp-241 and Leu-282, which includes many leucine residues in an alternating pattern. An amino acid sequence repeat is also located within that segment. Both of these patterns are present in human SBP and in the androgen-binding protein of rat epididymis. The sequence data indicate that the previously reported microheterogeneity of rabbit SBP in sodium dodecyl sulfate-polyacrylamide gel electrophoresis reflects variants generated by differential glycosylation of the monomer rather than different gene products. Seventy-nine percent of the amino acids of rabbit SBP are identical to those of human SBP; rabbit SBP thus joins human SBP and rat androgen-binding protein in one gene family that is distinct from the steroid hormone receptor superfamily. It appears that the problem of binding sex steroid hormones has been solved independently in two different gene families that contain completely different steroid-binding domains. Since the nonhomologous steroid-binding domains of both families of proteins recognize essentially the same steroid structure, it will be interesting to determine the structural basis of the two different protein designs that lead to similar steroid-binding specificity.
利用质谱分析和埃德曼降解技术的互补组合，确定了兔血清中特异性结合睾酮和5α-二氢睾酮的性类固醇结合蛋白（SBP或SHBG）的氨基酸序列。该同二聚体蛋白的单体单元是一条由367个氨基酸残基组成的单链糖肽，在Asn-345和Asn-361处有N-连接的寡糖侧链，二硫键将Cys-158与Cys-182以及Cys-327与Cys-355相连。根据序列计算的单体多肽分子量为39,769。包含9%碳水化合物的同二聚体分子量为87,404。该序列在Trp-241和Leu-282之间包含一个相对疏水的片段，其中有许多亮氨酸残基以交替模式排列。一个氨基酸序列重复也位于该片段内。这两种模式在人SBP和大鼠附睾雄激素结合蛋白中都存在。序列数据表明，先前报道的兔SBP在十二烷基硫酸钠-聚丙烯酰胺凝胶电泳中的微不均一性反映的是单体糖基化差异产生的变体，而非不同的基因产物。兔SBP 79%的氨基酸与人SBP相同；因此，兔SBP与人类SBP和大鼠雄激素结合蛋白属于同一个不同于类固醇激素受体超家族的基因家族。似乎在两个完全不同的包含不同类固醇结合结构域的基因家族中，独立解决了结合性类固醇激素的问题。由于这两个蛋白质家族的非同源类固醇结合结构域识别基本相同的类固醇结构，确定导致相似类固醇结合特异性的两种不同蛋白质设计的结构基础将很有趣。
