Human Basal Tear Peptidome Characterization by CID, HCD, and ETD Followed by in Silico and in Vitro Analyses for Antimicrobial Peptide Identification
Human Basal Tear Peptidome Characterization by CID, HCD, and ETD Followed by in Silico and in Vitro Analyses for Antimicrobial Peptide Identification
复制标题
DOI：
10.1021/acs.jproteome.5b00179
复制
发表时间：
2015-06-01
影响因子：
4.4
通讯作者：
Elortza, Felix
中科院分区：
文献类型：
--
作者：
Azkargorta, Mikel;Soria, Javier;Elortza, Felix
内源性肽是生物过程分析中的重要靶点。泪膜含有由泪管粘膜细胞释放的蛋白质和肽，包括参与保护免受外源性病原体侵害的抗微生物肽;然而，泪液的肽含量仍然表征不佳。我们分析了从人类基底泪液中分离的天然存在的肽。内源性肽的质谱分析存在许多缺点，包括大小异质性和不可预测的片段化模式等。因此，使用CID、ETD和HCD方法表征泪液肽含量。还评价了DMSO作为色谱溶剂添加剂的贡献。我们确定了157，131，和122肽使用CID-，ETD-，和HCD为基础的方法，分别。总共鉴定了234种不同的肽，从而产生了迄今为止最大的内源性泪液肽数据集。计算机模拟进行的抗微生物活性预测分析揭示了不同的推定抗微生物肽。两个细胞外糖蛋白lacritin肽从头合成，并在体外证实其抗菌活性。我们的研究结果表明，使用不同的片段化方法的内源性肽的分析的好处，并提供了一个有用的方法，发现具有抗菌活性的肽。
Endogenous peptides are valuable targets in the analysis of biological processes. The tear film contains proteins and peptides released by the tear duct mucosal cells, including antimicrobial peptides involved in the protection against exogenous pathogens; however, the peptide content of the tear liquid remains poorly characterized. We analyzed naturally occurring peptides isolated from human basal tears. Mass spectrometry analysis of endogenous peptides presents a number of drawbacks, including size heterogeneity and nonpredictable fragmentation patterns, among others. Therefore, CID, ETD, and HCD methods were used for the characterization of the tear peptide content. The contribution of DMSO as an additive of the chromatographic solvents was also evaluated. We identified 157, 131, and 122 peptides using CID-, ETD-, and HCD-based methods, respectively. Altogether, 234 different peptides were identified, leading to the generation of the biggest data set of endogenous tear peptides to date. The antimicrobial activity prediction analysis performed in silico revealed different putative antimicrobial peptides. Two of the extracellular glycoprotein lacritin peptides were de novo synthesized, and their antimicrobial activity was confirmed in vitro. Our findings demonstrate the benefits of using different fragmentation methods for the analysis of endogenous peptides and provide a useful approach for the discovery of peptides with antimicrobial activity.