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N. M. Kalariya, K. V. Ramana, S. K. Srivastava, F. J. G. M. van Kuijk; Posttranslational Modification of Proteins by Lutein Derived Aldehydes. Invest. Ophthalmol. Vis. Sci. 2008;49(13):4968.
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Lutein, commonly used as antioxidants to treat age-related macular degeneration (AMD), is also known to generate various cytotoxic aldehydes and cause cytotoxicity. Therefore, our aim was to understand how such aldehyde products could contribute to AMD instead of preventing it.
Proteins such as bovine serum albumin (BSA), α-crystallin and glucose-6-phosphate dehydrogenase (G6PDH) were incubated with lutein -derived aldehydes (LDAs) to examine posttranslational modification by performing gel electrophoresis, high performance liquid chromatography (HPLC) and mass spectroscopy (MALDI-TOF) and finally by amino acid analysis. The regulation of enzyme activity upon modification with LDAs was also determined by spectrophotometrically. The changes in carbonyl contents were quantified by carbonyl assay using 2,4-dinitrophenylhydrazine (2,4-DNP).
MALDI-TOF and SDS-PAGE revealed increase in the protein mass corresponding to protein adduct formation with LDAs. The carbonyl assay using 2,4-DNP suggests the possible involvement of Schiff’s base reaction between protein and LDAs. Amino acid analysis revealed possible interaction of LDAs with lysine amino acid residues of proteins. We also found that LDAs can form adducts with proteins by Michael’s adduct. Further formation of LDAs adducts with G6PDH significantly lost its catalytical activity in a time and dose dependent manner.
Our results suggest that LDAs could regulate protein function by posttranslationally by interacting with amino and sulfydryl group containing amino acids by Schiff’s and Michael’s reactions, respectively. Thus formation of LDAs adducts with proteins could inactivate the key enzymes responsible for maintaining cell homeostasis thereby causing cytotoxicity.
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