Fung Kim Y C, Morris Carol, Sathe Sonal, Sack Robert, Duncan Mark W
Biochemical Mass Spectrometry Facility University of Colorado Health Sciences Center Denver, CO, USA.
Proteomics. 2004 Dec;4(12):3953-9. doi: 10.1002/pmic.200300849.
The tear film is complex and is rich in both peptides and proteins. Physiological factors have been shown to alter the balance of the protein components in the tear film, however, little is known of the precise stimuli that initiate these changes, or their nature and extent. Attention has been directed at the role of tear proteins in the protection of the external ocular surface, and their potential role in the pathogenesis of inflammatory and autoimmune diseases, but few lacrimal-specific proteins have been identified and demonstrated to offer a protective function at the ocular surface. The biological importance of proline-rich proteins is uncertain, although there is some evidence to indicate a potential antimicrobial function for these proteins in saliva. Despite the detection of mRNA for proline-rich proteins in lacrimal gland, the translated protein product has not been detected in tear fluid. In this study we investigate the presence of proline-rich proteins in the tear film. Human reflex tear fluid was examined by matrix-assisted laser desorption/ionization-time of flight mass spectrometry directly, and following size exclusion high performance liquid chromatography. This revealed significant levels of a truncated form of lacrimal proline-rich protein, and a series of peptides derived the C-terminus of this protein. None of these had previously been identified in tear. Our study highlights the dangers inherent in proteomic strategies that assign an identity to a protein based on limited coverage of tryptic peptides.
泪膜结构复杂，富含多种肽和蛋白质。生理因素可改变泪膜中蛋白质成分的平衡，然而，引发这些变化的精确刺激因素及其性质和程度却鲜为人知。人们已将注意力投向泪液蛋白质在保护眼表方面的作用，以及它们在炎症和自身免疫性疾病发病机制中的潜在作用，但目前仅鉴定出少数泪腺特异性蛋白质，并证实其在眼表具有保护功能。富含脯氨酸的蛋白质的生物学重要性尚不确定，尽管有证据表明这些蛋白质在唾液中可能具有抗菌功能。尽管在泪腺中检测到了富含脯氨酸蛋白质的信使核糖核酸，但在泪液中尚未检测到其翻译后的蛋白质产物。在本研究中，我们对泪膜中富含脯氨酸的蛋白质进行了研究。采用基质辅助激光解吸/电离飞行时间质谱直接检测以及排阻高效液相色谱法对人类反射性泪液进行了检测。结果显示，泪腺富含脯氨酸蛋白质的一种截短形式以及一系列源自该蛋白质C末端的肽含量显著。此前在泪液中均未鉴定出这些成分。我们的研究强调了蛋白质组学策略中存在的风险，即基于胰蛋白酶肽段的有限覆盖度来确定蛋白质的身份。
