Ribosomal Synthesis of Peptides with Unnatural Building Blocks

描述（由申请人提供）：我们将尝试使用我们的非天然氨基酸诱变方法对含有多种非天然氨基酸的环状和线性肽库进行遗传编码。这些肽文库将在细菌中细胞内产生，并以噬菌体展示的形式产生，以分别允许体内选择和基于展示的亲和淘选。这种策略将使分子结构的直接遗传编码、随机或定向诱变、选择和生物扩增成为可能，同时允许化学多样性超出典型的20个氨基酸库所能提供的范围。将非天然氨基酸整合到遗传编码的肽文库中应有助于进化实验和产生具有新生物活性的肽，这些肽最终可能会发现治疗用途。这将通过以下具体目标实现：1.制备和掺入具有独特反应性和具有能够靶向参与疾病状态的特定蛋白质（例如，丝氨酸蛋白酶、金属蛋白酶等）2.利用内含肽为基础的胞内环肽合成方法结合非天然氨基酸技术构建新型环肽文库并筛选特异性酶抑制剂3.使用噬菌体展示系统来产生和筛选含有非天然氨基酸的肽文库以用于特异性蛋白质靶点的抑制剂。

DESCRIPTION (provided by applicant): We will attempt to use our unnatural amino acid mutagenesis methodology to genetically encode libraries of cyclic and linear peptides containing a diverse array of unnatural amino acids. These peptide libraries will be generated both intracellularly in bacteria and in a phage-displayed format to allow in vivo selections and display-based affinity panning, respectively. This strategy will enable direct genetic encoding of molecular structure, random or directed mutagenesis, selection, and biological amplification, while at the same time allowing chemical diversity beyond what the canonical 20 amino acid repertoire can offer. The integration of unnatural amino acids into genetically encoded peptide libraries should facilitate both evolutionary experiments and the generation of peptides with novel biological activities that may ultimately find therapeutic use. This will be achieved through the following specific aims: 1. The preparation and incorporation of novel unnatural amino acids with unique reactivities and with functional groups capable of targeting specific proteins involved in disease states (e.g., serine proteases, metalloproteases, etc.) 2. The use of intein-based intracellular cyclic peptide synthesis methodologies together with unnatural amino acid technology to generate novel cyclic peptide libraries and select specific enzyme inhibitors 3. The use of phage display systems to generate and screen libraries of peptides containing unnatural amino acids for inhibitors of specific protein targets.