Zot H G, Bhaskara V, Liu L
Department of Biology, Eastern Michigan University, Ypsilanti, Michigan, 48197, USA.
Arch Biochem Biophys. 2000 Mar 1;375(1):161-4. doi: 10.1006/abbi.1999.1648.
The domain organization of Acanthamoeba myosin-I, an oligomodular motor protein, includes a potentially important protein interaction module that is mostly uncharacterized. The Src homology 3, SH3, domain of myosin-I binds Acan125, a protein containing at least two consensus ligand binding domains: C-terminal SH3 binding motifs (PXXP) and N-terminal leucine-rich repeats. We report the first affinities determined for an SH3 domain of any myosin, namely, K(d) = 7 microM for a 21-residue synthetic peptide based on the PXXP domain sequence and K(d) = 0.15 microM for the PXXP domain included in the C-terminus of Acan125. These values are consistent with affinities reported for peptides and proteins that associate with SH3. By deletional analysis we show that only the PXXP domain is required for Acan125 to interact with the SH3 domain of Acanthamoeba myosin-IC (AmyoC(SH3)). The synthetic peptide described above at a concentration near the K(d) for SH3 binding blocked the interaction between native AmyoC and Acan125, mapping the interaction to the PXXP domain of Acan125 and the SH3 domain of myosin-I. These results are consistent with prototypical SH3 binding and suggest that a PXXP module is both necessary and sufficient to interact with an SH3 module of myosin-I.
棘阿米巴肌球蛋白-I是一种寡聚模块化运动蛋白，其结构域组织包括一个潜在的重要蛋白相互作用模块，而该模块大多尚未得到充分研究。肌球蛋白-I的Src同源3（SH3）结构域与Acan125结合，Acan125是一种至少含有两个共有配体结合结构域的蛋白：C端SH3结合基序（PXXP）和N端富含亮氨酸重复序列。我们报道了首个针对任何肌球蛋白的SH3结构域测定的亲和力，即基于PXXP结构域序列的21个残基合成肽的解离常数K(d)=7 microM，以及Acan125 C端包含的PXXP结构域的K(d)=0.15 microM。这些值与报道的与SH3相关的肽和蛋白的亲和力一致。通过缺失分析，我们表明Acan125与棘阿米巴肌球蛋白-IC（AmyoC(SH3)）的SH3结构域相互作用仅需要PXXP结构域。上述浓度接近SH3结合K(d)的合成肽阻断了天然AmyoC与Acan125之间的相互作用，将这种相互作用定位到Acan125的PXXP结构域和肌球蛋白-I的SH3结构域。这些结果与典型的SH3结合一致，表明一个PXXP模块对于与肌球蛋白-I的SH3模块相互作用既是必要的也是充分的。
