Zarn J A, Lüthi C, Giger R J, Sigrist A, Humbel R E
Department of Biochemistry, University of Zürich, Switzerland.
Eur J Biochem. 1992 Dec 15;210(3):665-9. doi: 10.1111/j.1432-1033.1992.tb17467.x.
A mutant of human insulin-like growth factor II (IGF II) was constructed by site-directed mutagenesis: the nucleotides coding for Ser33 and Ser39 were changed to yield Arg and Lys, respectively, thus creating two pairs of basic residues, Arg-Arg and Lys-Arg, as flanking sequences of the remaining C domain. [Arg33, Lys39]IGF II was expressed in NIH-3T3 cells as a processed two-chain peptide with a deletion of amino acid residues 37-40 and crosslinked by three disulfide bonds. This des(37-40)[Arg33]IGF II showed 3.6-fold and 7.4-fold reduced affinities to the type 1 and type 2 IGF receptor overexpressing cells, respectively, whereas the thymidine incorporation potency was the same as that of wild-type IGF II. We speculate that the discrepancy between the reduced binding to the type 1 IGF receptor and the full thymidine incorporation potency is due to the 6.1-fold reduced affinity of the expressed mutant to the co-expressed IGF binding protein 3 (IGFBP-3). The results suggest that des(37-40)[Arg33]IGF II assumes a conformation very similar to IGF II, and that the entire length of the C domain is not essential for biological activity.
通过定点诱变构建了人胰岛素样生长因子II（IGF II）的突变体：编码Ser33和Ser39的核苷酸发生改变，分别产生Arg和Lys，从而形成两对碱性残基，即Arg-Arg和Lys-Arg，作为剩余C结构域的侧翼序列。[Arg33，Lys39]IGF II在NIH-3T3细胞中表达为一种加工后的双链肽，缺失氨基酸残基37-40，并通过三个二硫键交联。这种缺失（37-40）[Arg33]IGF II对过表达1型和2型IGF受体的细胞的亲和力分别降低了3.6倍和7.4倍，而胸苷掺入能力与野生型IGF II相同。我们推测，与1型IGF受体结合减少和胸苷掺入能力完全之间的差异是由于所表达的突变体与共表达的IGF结合蛋白3（IGFBP-3）的亲和力降低了6.1倍。结果表明，缺失（37-40）[Arg33]IGF II呈现出与IGF II非常相似的构象，并且C结构域的全长对于生物活性不是必需的。
