Institut Químic de Sarrià (IQS), Universitat Ramon Llull, Barcelona, Spain.
Research School of Biology, The Australian National University, Canberra, Australia.
Methods Mol Biol. 2023;2676:117-129. doi: 10.1007/978-1-0716-3251-2_8.
Phage display facilitates the evolution of peptides and proteins for affinity selection against targets, but it is mostly limited to the chemical diversity provided by the naturally encoded amino acids. The combination of phage display with genetic code expansion allows the incorporation of noncanonical amino acids (ncAAs) into proteins expressed on the phage. In this method, we describe incorporation of one or two ncAAs in a single-chain fragment variable (scFv) antibody in response to amber or quadruplet codon. We take advantage of the pyrrolysyl-tRNA synthetase/tRNA pair to incorporate a lysine derivative and an orthogonal tyrosyl-tRNA synthetase/tRNA pair to incorporate a phenylalanine derivative. The encoding of novel chemical functionalities and building blocks in proteins displayed on phage provides the foundation for further phage display applications in fields such as imaging, protein targeting, and the production of new materials.
噬菌体展示技术促进了肽和蛋白质的进化，以针对目标进行亲和选择，但它主要限于天然编码氨基酸提供的化学多样性。噬菌体展示与遗传密码扩展的结合允许将非天然氨基酸（ncAAs）掺入到噬菌体上表达的蛋白质中。在这种方法中，我们描述了在单链片段可变区（scFv）抗体中响应琥珀酰或四联体密码子掺入一个或两个 ncAAs。我们利用吡咯赖氨酸-tRNA 合成酶/tRNA 对掺入赖氨酸衍生物，并用正交的酪氨酸-tRNA 合成酶/tRNA 对掺入苯丙氨酸衍生物。在噬菌体上展示的蛋白质中编码新型化学功能和构建块为噬菌体展示在成像、蛋白质靶向和新材料生产等领域的进一步应用奠定了基础。
