Bio-Inspired Design and Potential Biomedical Applications of a Novel Class of High-Affinity Peptides
Bio-Inspired Design and Potential Biomedical Applications of a Novel Class of High-Affinity Peptides
复制标题
DOI：
10.1002/anie.201107894
复制
发表时间：
2012-01-01
影响因子：
16.6
通讯作者：
Jon, Sangyong
中科院分区：
文献类型：
--
作者：
Kim, Sunghyun;Kim, Daejin;Jon, Sangyong
抗体由于其固有的高亲和力和对多种靶点的特异性，已广泛应用于生物制药和生物医学领域然而，由于它们的体积大，不希望的效应功能，免疫原性，在哺乳动物细胞中昂贵的重组生产以及复杂的知识产权壁垒（版税叠加），导致组织穿透性差，导致研究人员寻找抗体的替代品基于蛋白支架的亲和分子[3]和寡核苷酸或基于rna的适体[4]最近作为新的高亲和分子出现，在诊断和治疗中确实显示出潜在的效用。这类高亲和分子的一个共同特点是具有三维折叠结构，有利于通过大的识别界面结合靶标，从而使靶标结合紧密，特异性高。具有小分子质量的高亲和性分子比大的高亲和性分子表现出快速的外渗和更高的组织渗透，可以用作癌症诊断[6]或治疗药物[7]然而，迄今为止，很少有报道描述了基于肽支架的亲和分子的发展，可能是因为肽很难形成强大的预组织结构。结蛋白家族[8]和层聚物[9]是从天然存在的蛋白结构域工程化的肽支架；可以从这两个家族中选择具有微摩尔[10]或纳摩尔亲和[11]的紧密肽结合物。然而，多个二硫键参与稳定结蛋白支架。因此，结构确定是必要的，以确保假设正确的折叠，这限制了产品开发的速度支聚体不是具有确定的单一结构框架的肽支架；它们是由来自国外的各种自然发生的结构框架组成的库。因此，必须解决选定的hit的结构以找出特定的支架，并且由于层聚体的起源，存在潜在的免疫原性问题。[9]据我们所知，目前还没有报道描述人工肽支架可以作为高亲和力肽的一般来源，并对纳摩尔范围内的各种生物靶点表现出亲和力。人造肽支架必须满足以下设计标准：1)能够形成二级结构；2)其预组织结构应稳定、稳健；3)它应该有足够数量的可变（随机）氨基酸来创造多样性；4)其可变区域应尽量不影响其二级结构。在此，受碱性亮氨酸拉链蛋白（bZIP）结构的启发，我们合理设计了基于人工肽支架的新型亲和分子，这些分子形成了强大的预组织结构，能够以高亲和性和特异性结合靶标。bZIP蛋白在所有真核生物中都通过高亲和力（低纳摩尔）、序列特异性识别独特的DNA基序发挥转录调节作用。同源或异二聚体bZIP蛋白有一个毗邻七亮氨酸重复序列的基本区域（图1a）。上亮氨酸拉链区作为一个支架，维持一个独特的开口结构，通过这个结构，可变的基本区域被用来识别DNA位点利用这些特征，我们设计了新的人工高亲和力肽配体，我们称之为“aptides”，来自适配体样肽。肽包括一个稳定支架和两个靶标结合区（图1a）。支架由一个小的（12个氨基酸）但高度稳定的色氨酸拉链（trpzip; Tm= 728C）组成。
Antibodies have been widely used in a range of biopharmaceutical and biomedical applications due to their intrinsic high affinity and specificity toward various targets.[1] However, poor tissue penetration owing to their large size, undesired effector functions, immunogenicity, costly recombinant production in mammalian cells, and complex intellectual property barriers (royalty stacking) have led researchers to seek alternatives to antibodies.[2] Protein-scaffold-based affinity molecules [3] and oligo DNA or RNA-based aptamers [4] have recently emerged as novel high-affinity molecules that have indeed demonstrated potential utility in diagnosis and therapy. A common feature of such high-affinity molecules is that they possess three-dimensional folded structures that facilitate target binding through a large recognition interface, resulting in tight target binding with high specificity. Highaffinity molecules with small molecular mass exhibit rapid extravasation and higher tissue penetration than bigger highaffinity molecules [5] and can be used as cancer diagnostics [6] or therapeutics.[7] To date, however, few reports have described the development of peptide-scaffold-based affinity molecules, presumably because it is difficult for peptides to form robust pre-organized structures. The knottin family [8] and phylomers [9] are peptide scaffolds engineered from naturally occurring protein domains; tight peptide binders with micromolar [10] or nanomolar affinities [11] can be selected from these two families. However, multiple disulfide bonds are involved in stabilizing knottin scaffolds. Thus, structural determination is necessary to ensure assumption of the correct fold, which limits the speed of product development.[12] Phylomers are not peptide scaffolds with a defined, single structural framework; they rather consist of a library of diverse naturally occurring structural frameworks derived from foreign sources. Therefore, the structures of selected hits must be solved to figure out the specific scaffold, and there is a concern on potential immunogenicity because of the origin of phylomers.[9]To our knowledge, no reports have described artificial peptide scaffolds that can be used as a general source of highaffinity peptides and that exhibit affinities toward a variety of biological targets in the nanomolar range. An artificial peptide scaffold must satisfy the following design criteria: 1) it should be able to form a secondary structure; 2) its preorganized structure should be stable and robust; 3) it should have a sufficient number of variable (randomizable) amino acids to create diversity; and 4) its variable regions should minimally affect its secondary structure. Herein, inspired by the structure of basic leucine zipper (bZIP) proteins, which function as transcriptional regulators in all eukaryotes through high-affinity (ca. low nanomolar), sequence-specific recognition of unique DNA motifs, we rationally designed novel artificial peptide-scaffold-based affinity molecules that form robust pre-organized structures and are capable of binding targets with high affinity and specificity. Homo-or heterodimeric bZIP proteins have a basic region that abuts a sequence of hepta-leucine repeats (Figure1a). The upper leucine-zipper region serves as a scaffold that maintains a unique open-mouthed structure through which variable basic regions are used to recognize DNA sites.[13] Exploiting these features, we designed new artificial high-affinity peptide ligands, which we have termed “aptides”, from aptamer-like peptides. An aptide comprises a stabilizing scaffold and two target-binding regions (Figure 1a). The scaffold consists of a small (12 amino acids) but highly stable tryptophan zipper (trpzip; Tm= 728C) that …