1 research outputs found
Human Basal Tear Peptidome Characterization by CID, HCD, and ETD Followed by in Silico and in Vitro Analyses for Antimicrobial Peptide Identification
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