Yang Xu, Dong Xiangchao, Zhang Kai, Yang Fangfang, Guo Zhenchang
College of Chemistry, Collaborative Innovation Center of Chemical Science and Engineering (Tianjin), Nankai University, Tianjin 300071, China.
J Mater Chem B. 2016 Feb 7;4(5):920-928. doi: 10.1039/c5tb02620b. Epub 2016 Jan 8.
Lysine acetylation is a widespread protein post-translational modification (PTM) that plays a central role in diverse physiological processes. The study on the scope and pattern of lysine acetylation is an important subject in the proteomic research. However, identification of lysine acetylation from biological sources is a great challenge due to the low abundance in the massive background of unmodified proteins and the dynamic fashion of the modifications. In this research, a novel molecularly imprinted polymer (MIP) with high affinity for peptides containing acetylated lysine (Kac) has been synthesized by the combination of an epitope and surface-confined imprinting strategy. A dipeptide: KacA, containing acetylated lysine and alanine residues, was used as the template and immobilized on the sacrificial silica support. After hierarchical imprinting and removal of silica, the surface-confined cavities were created on the resulting KacA-MIP material. The equilibrium binding and HPLC experiments demonstrated that the KacA-MIP has good selectivity and epitope affinity. It can differentiate Lys-acetylated peptides (Kac-peptides) from their native structures and has higher affinity for Kac-peptides with different sequences. The selectivity of the MIP was also proved by its ability in the extraction of Kac-peptides from spiked histone digest and by its enrichment performance in the whole cell lysates. The study developed a method of MIP preparation with affinity for PTM peptide based on recognition of peptide side chains. It also indicated that the MIP has potential to be used as an antibody mimic in the PTM analysis.
赖氨酸乙酰化是一种广泛存在的蛋白质翻译后修饰（PTM），在多种生理过程中发挥着核心作用。赖氨酸乙酰化的范围和模式研究是蛋白质组学研究中的一个重要课题。然而，由于未修饰蛋白质的大量背景中丰度较低以及修饰的动态特性，从生物来源鉴定赖氨酸乙酰化是一项巨大挑战。在本研究中，通过表位与表面受限印迹策略相结合，合成了一种对含乙酰化赖氨酸（Kac）的肽具有高亲和力的新型分子印迹聚合物（MIP）。一种含有乙酰化赖氨酸和丙氨酸残基的二肽：KacA，被用作模板并固定在牺牲性二氧化硅载体上。经过分级印迹和二氧化硅去除后，在所得的KacA-MIP材料上形成了表面受限的空腔。平衡结合和高效液相色谱实验表明，KacA-MIP具有良好的选择性和表位亲和力。它可以区分赖氨酸乙酰化肽（Kac-肽）与其天然结构，并且对不同序列的Kac-肽具有更高的亲和力。MIP的选择性还通过其从加标的组蛋白消化物中提取Kac-肽的能力及其在全细胞裂解物中的富集性能得到证明。该研究开发了一种基于肽侧链识别的对PTM肽具有亲和力的MIP制备方法。它还表明MIP在PTM分析中具有作为抗体模拟物的潜力。
