MALDI Imaging Mass Spectrometry Spatially Maps Age-Related Deamidation and Truncation of Human Lens Aquaporin-0.

Wenke JL, Rose KL, Spraggins JM, Schey KL
Invest Ophthalmol Vis Sci. 2015 56 (12): 7398-405

PMID: 26574799 · PMCID: PMC4675204 · DOI:10.1167/iovs.15-18117

PURPOSE - To spatially map human lens Aquaporin-0 (AQP0) protein modifications, including lipidation, truncation, and deamidation, from birth through middle age using matrix-assisted laser desorption ionization (MALDI) imaging mass spectrometry (IMS).

METHODS - Human lens sections were water-washed to facilitate detection of membrane protein AQP0. We acquired MALDI images from eight human lenses ranging in age from 2 months to 63 years. In situ tryptic digestion was used to generate peptides of AQP0 and peptide images were acquired on a 15T Fourier transform ion cyclotron resonance (FTICR) mass spectrometer. Peptide extracts were analyzed by liquid chromatography-tandem mass spectrometry (LC-MS/MS) and database searched to identify peptides observed in MALDI imaging experiments.

RESULTS - Unmodified, truncated, and fatty acid-acylated forms of AQP0 were detected in protein imaging experiments. Full-length AQP0 was fatty acid acylated in the core and cortex of young (2- and 4-month) lenses. Acylated and unmodified AQP0 were C-terminally truncated in older lens cores. Deamidated tryptic peptides (+0.9847 Da) were mass resolved from unmodified peptides by FTICR MS. Peptide images revealed differential localization of un-, singly-, and doubly-deamidated AQP0 C-terminal peptide (239-263). Deamidation was present at 4 months and increases with age. Liquid chromatography-MS/MS results indicated N246 undergoes deamidation more rapidly than N259.

CONCLUSIONS - Results indicated AQP0 fatty acid acylation and deamidation occur during early development. Progressive age-related AQP0 processing, including deamidation and truncation, was mapped in human lenses as a function of age. The localization of these modified AQP0 forms suggests where AQP0 functions may change throughout lens development and aging.

MeSH Terms (15)

Adult Aging Aquaporins Cataract Eye Proteins Female Fourier Analysis Humans Lens, Crystalline Male Middle Aged Protein Processing, Post-Translational Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization Tandem Mass Spectrometry Young Adult

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