Esterification of protein carboxylic acids results in a myriad of biological functions. Such modifications generally are reversible. As such, the esterification states are often dynamic. In order to build a comprehensive view of protein carboxylic esterification, techniques for assessing the extent or patterns of these modifications are necessary. However, the current methods, which are heavily dependent on radioactive labeling, do not provide the specificity and sensitivity required. To fill up this important gap, we propose herein to develop two general methods to detect and quantify protein carboxylic esters.
Specific Aim 1 is to devise bi-functional derivatization reagents for carboxylic esters. Our strategy involves an initial acyl transfer between esters and strong nucleophites, such as hydrazine; and the subsequent coupling to various tags that is catalyzed by click chemistry.
Specific Aim 2 is to synthesize AdoMet analogs to covalently label carboxylic groups that are subject to methylation. This strategy will be especially valuable in detecting carboxylic esters that only exist transiently. ? ?
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| Alfaro, Joshua F; Gillies, Laura A; Sun, He G et al. (2008) Chemo-enzymatic detection of protein isoaspartate using protein isoaspartate methyltransferase and hydrazine trapping. Anal Chem 80:3882-9 |
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