In-gel protein digestion using acidic methanol produces a highly selective methylation of glutamic 1 acid residues.

Lozano-Prieto, M.; Camafeita, E.; Jorge, I.; Laguillo-Gomez, A.; Barrero-Rodriguez, R.; Devesa, C. A.; Pertusa, C.; Calvo, E.; Sanchez-Madrid, F.; Vazquez, J.; Martin-Cofreces, N. B.

doi:10.1101/2024.03.15.585290

bioRxiv · 10.1101/2024.03.15.585290

In-gel protein digestion using acidic methanol produces a highly selective methylation of glutamic 1 acid residues.

Lozano-Prieto, M.·Camafeita, E.·Jorge, I.·Laguillo-Gomez, A.·Barrero-Rodriguez, R.·Devesa, C. A.·Pertusa, C.·Calvo, E.·Sanchez-Madrid, F.·Vazquez, J.·Martin-Cofreces, N. B.

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Abstract

Mass-tolerant open search methods allow the high-throughput analysis of modified peptides by mass spectrometry. These techniques have paved the way to unbiased analysis of post-translational modifications (PTMs) in biological contexts, as well as of chemical modifications produced during the manipulation of protein samples. In this work, we have analyzed in-depth a wide variety of samples of different biological origin, including cells, extracellular vesicles, secretomes, centrosomes and tissue preparations, using Comet-ReCom, a recently improved version of the open search engine Comet-PTM. Our results demonstrate that glutamic acid residues undergo intensive methyl esterification when protein digestion is performed using in-gel techniques, but not using gel-free approaches. This effect was highly specific to Glu and was not found for other methylable residues such as Asp.

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Lozano-Prieto, M., Camafeita, E., Jorge, I., Laguillo-Gomez, A., Barrero-Rodriguez, R., Devesa, C. A., Pertusa, C., Calvo, E., Sanchez-Madrid, F., Vazquez, J., Martin-Cofreces, N. B.. 2024-03-17. In-gel protein digestion using acidic methanol produces a highly selective methylation of glutamic 1 acid residues.. https://doi.org/10.1101/2024.03.15.585290

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