Department of Biotechnology, Friedrich-Alexander University Erlangen-Nuremberg, 91054 Erlangen, Germany.
Single-Molecule Microscopy Group, Jena University Hospital, Friedrich-Schiller University Jena, 07743 Jena, Germany.
Proc Natl Acad Sci U S A. 2018 May 29;115(22):5744-5749. doi: 10.1073/pnas.1716666115. Epub 2018 May 14.
The allosteric interplay between distant functional sites present in a single protein provides for one of the most important regulatory mechanisms in biological systems. While the design of ligand-binding sites into proteins remains challenging, this holds even truer for the coupling of a newly engineered binding site to an allosteric mechanism that regulates the ligand affinity. Here it is shown how computational design algorithms enabled the introduction of doxycycline- and doxorubicin-binding sites into the serine proteinase inhibitor (serpin) family member α1-antichymotrypsin. Further engineering allowed exploitation of the proteinase-triggered serpin-typical S-to-R transition to modulate the ligand affinities. These design variants follow strategies observed in naturally occurring plasma globulins that allow for the targeted delivery of hormones in the blood. By analogy, we propose that the variants described in the present study could be further developed to allow for the delivery of the antibiotic doxycycline and the anticancer compound doxorubicin to tissues/locations that express specific proteinases, such as bacterial infection sites or tumor cells secreting matrix metalloproteinases.
在单个蛋白质中存在的远距离功能位点的变构相互作用提供了生物系统中最重要的调节机制之一。虽然将配体结合位点设计到蛋白质中仍然具有挑战性，但对于将新设计的结合位点与调节配体亲和力的变构机制相耦合来说，更是如此。本文展示了如何使用计算设计算法将强力霉素和阿霉素结合位点引入丝氨酸蛋白酶抑制剂（丝氨酸蛋白酶抑制剂）家族成员α1-抗胰蛋白酶。进一步的工程设计允许利用蛋白酶触发的丝氨酸蛋白酶典型的 S 到 R 转变来调节配体亲和力。这些设计变体遵循在天然存在的血浆球蛋白中观察到的策略，允许在血液中靶向递送激素。通过类比，我们提出在本研究中描述的变体可以进一步开发，以允许将抗生素强力霉素和抗癌化合物阿霉素递送至表达特定蛋白酶的组织/位置，例如细菌感染部位或分泌基质金属蛋白酶的肿瘤细胞。
