info:eu-repo/semantics/article
Online electrospray ionization mass spectrometric monitoring of protease-catalyzed reactions in real time
Fecha
2012-04Registro en:
Yu, Zhan; Chen, Lee Chuin; Mandal, Mridul Kanti; Nonami, Hiroshi; Erra Balsells, Rosa; et al.; Online electrospray ionization mass spectrometric monitoring of protease-catalyzed reactions in real time; Springer; Journal of The American Society for Mass Spectrometry; 23; 4; 4-2012; 728-735
1044-0305
CONICET Digital
CONICET
Autor
Yu, Zhan
Chen, Lee Chuin
Mandal, Mridul Kanti
Nonami, Hiroshi
Erra Balsells, Rosa
Hiraoka, Kenzo
Resumen
Although there are a lot of well established methods for monitoring enzyme-catalyzed reactions, most of them are based on changes in spectroscopic properties during the conversion of substrates to products. However, reactions without optical changes are common, which are inapplicable to these spectroscopic methods. As an alternative technique for enzymologic research, mass spectrometry (MS) is favored due to its specificity, sensitivity, and the ability to obtain stoichiometric information. In this work, probe electrospray ionization (PESI) source coupled with a time of flight mass spectrometer was employed to monitor some typical proteasecatalyzed reactions, including pepsinolysis and trypsinolysis of cytochrome c in real time. Due to the high electrical conductivity of each reaction system, corona discharges are likely to occur, which would decrease intensities of mass spectrometric signals. An ultra-fine sampling probe and an auxiliary vapor spray were adopted to prevent corona discharges. Experimental results from peptic and tryptic digestions of cytochrome c showed different and characteristic catalytic pathways. With the data presented in this study, PESI-MS can be considered as a potential tool for real-time monitoring of enzymatic reactions because of its simplicity in instrumental configuration, wide applicability under harsh conditions, and flexibility in combination with other techniques.