Ptacek Jennifer B, Edwards Anne P B, Freeman-Cook Lisa L, DiMaio Daniel
Department of Genetics and Therapeutic Radiology, Yale University School of Medicine, 333 Cedar Street, New Haven, CT 06510, USA.
Proc Natl Acad Sci U S A. 2007 Jul 17;104(29):11945-50. doi: 10.1073/pnas.0704348104. Epub 2007 Jul 3.
We used proteins with randomized transmembrane (TM) domains to explore the role of hydrophobic amino acids in mediating specific interactions between transmembrane helices. The 44-aa bovine papillomavirus E5 protein, which binds to the TM domain of the PDGFbeta receptor (PDGFbetaR) was used as a scaffold to construct a library encoding small dimeric proteins with randomized, strictly hydrophobic TM domains, and proteins were selected that induced focus formation in mouse C127 cells by activating the PDGFbetaR. Analysis of these proteins identified a motif of two hydrophobic residues that, when inserted into a 17-residue polyleucine TM domain, generated a protein that activated the PDGFbetaR and transformed cells. In addition, we identified transforming proteins that activated the wild-type PDGFbetaR but did not activate a series of PDGFbetaR TM point mutants that were efficiently activated by the E5 protein, indicating that these proteins were more specific than the E5 protein. Our results implied that multiple van der Waals interactions distributed along the entire length of the TM domains were required for productive interaction between the PDGFbetaR and some small proteins lacking hydrophilic TM residues. Our results also suggested that excluding hydrophilic residues from small TM proteins and peptides is a strategy to increase the specificity of heteromeric TM helix-helix interactions.
我们使用具有随机跨膜（TM）结构域的蛋白质来探究疏水氨基酸在介导跨膜螺旋之间特异性相互作用中的作用。将与血小板衍生生长因子β受体（PDGFβR）的TM结构域结合的44个氨基酸的牛乳头瘤病毒E5蛋白用作支架，构建一个编码具有随机、严格疏水TM结构域的小二聚体蛋白的文库，并筛选出通过激活PDGFβR在小鼠C127细胞中诱导灶形成的蛋白质。对这些蛋白质的分析确定了一个由两个疏水残基组成的基序，当将其插入一个17个残基的聚亮氨酸TM结构域时，产生了一种激活PDGFβR并转化细胞的蛋白质。此外，我们鉴定出了激活野生型PDGFβR但不激活一系列被E5蛋白有效激活的PDGFβR TM点突变体的转化蛋白，这表明这些蛋白质比E5蛋白更具特异性。我们的结果表明，PDGFβR与一些缺乏亲水性TM残基的小蛋白质之间的有效相互作用需要沿着TM结构域的整个长度分布多个范德华相互作用。我们的结果还表明，从小的TM蛋白和肽中排除亲水性残基是提高异源TM螺旋-螺旋相互作用特异性的一种策略。
