Glycative Stress Research
Online ISSN : 2188-3610
Print ISSN : 2188-3602
ISSN-L : 2188-3610
Identification of acetaldehyde-modified lysine
BanSakakeKushidaManabeYoshiki YamaguchiNagai
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JOURNAL OPEN ACCESS

2021 Volume 8 Issue 4 Pages 201-210

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Abstract

Intermediate carbonyl compounds produced by abnormalities in glucose and lipid metabolism in vivo induce amino carbonyl reactions that modify proteins in a non-enzymatic manner. This reaction is further accelerated with the progression of lifestyle-related diseases, eventually leading to the formation of advanced glycation end-products (AGEs). Also, alcoholic beverages are consumed as a luxury item. Acetaldehyde (AA), which is produced as an intermediate metabolite of alcohol metabolism, has a highly reactive carbonyl group and may modify proteins in the same manner as AGEs are produced. In this study, we attempted to identify the structure of AA-modified amino acids (AA-A) and measure their amount by immunochemical methods using monoclonal antibodies in order to detect the modification of proteins by AA. The spleens of mice immunized with AA-keyhole limpet hemocyanin (AA-KLH) were fused with myeloma cells, and antibody-producing cells reactive with AA-bovine serum albumin (AA-BSA) were isolated. The amino acid analysis of AA-BSA showed that lysine (Lys) and histidine were highly modified. Focusing on Lys with amino groups on its side chain, AA-Lys was isolated from the product generated from Nα-benzyloxycarbonyl-L-lysine (Nα-Cbz-Lys) and AA by HPLC and the structure was analyzed by nuclear magnetic resonance (NMR) and mass spectrometry. As a result, m/z 385 and m/z 411 were detected, and the structure was identified by NMR. The reactivity of these three fractions with antibodies against monoclonal anti-AA-modified proteins was evaluated and the structures were analyzed. In addition, the three fractions in AA-modified BSA incubated in phosphate buffer (pH 7.4) at 37 oC for 7 days were evaluated by ELISA. As a result, the structures of the three fractions were identified, and the highest reactivity with antibodies was observed in fraction 2. These increased in a concentration-dependent manner with AA-modified BSA. These results indicate that three new AA-derived Lys structures have been identified and are formed under physiological conditions, and that this structure will be one of the tools to evaluate protein modification by AA.

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© 2021 Society for Glycative Stress Research
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