Characterization of lacrimal proline‐rich protein 4 ( PRR 4) in human tear proteome

This study was initiated considering the lack of comprehensive characteristics profile of PRR 4 in tears of healthy subjects. Therefore, detailed characterizations of PRR 4 from basal tears employing in‐gel and in‐solution digestions for MS systems are presented herein. First, pooled tear samples ( n = 10) were utilized to identify PRR 4‐rich region/spots in 1 DE /2 DE gels employing LC ‐ MALDI ‐ MS and 1 DE ‐ LC ‐ ESI ‐ LTQ ‐ O rbitrap‐ MS systems. PRR 4‐rich region and ten spots with vast polymorphisms ( M r : 17–30 kDa, p I : 3.0–6.6) were identified in 1 DE and 2 DE gels, respectively. In addition, combinations of four types of PTM s, which are methylation, acetylation, oxidation, and pyroglutamate formation, were identified in these ten PRR4 spots. Furthermore, a targeted data‐acquisition approach was utilized to identify PRR 4 isoforms in individual tear samples ( n = 61) by in‐solution digestion combined with a LC ‐ ESI ‐ LTQ ‐ O rbitrap‐ MS system. Importantly, a new PRR 4 isoform designated as PRR 4‐ N 3 in addition to PRR 4 (gi154448886) and p HL E 1 F 1 (gi1050983) was identified. Moreover, different combinations of these three PRR 4 isoforms identified in the individual tear samples could be categorized into six distinguished groups. Conclusively, these findings provide fundamental insight into the complex characteristics profile of PRR 4 isoforms and their PTM s in tears of healthy individuals.