Characterization of Unstable Intermediates and Oxidized Products Formed during Cyanogen Bromide Cleavage of Peptides and Proteins by Electrospray Mass Spectrometry

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• 作者：Xinyi Zhang ; Lieve Dillen ; Koen Vanhoutte ; Walter Van Dongen ; Eddy Esmans ; and Magda Claeys
• 刊名：Analytical Chemistry
• 出版年：1996
• 出版时间：October 1, 1996
• 年：1996
• 卷：68
• 期：19
• 页码：3422 - 3430
• 全文大小：491K
• 年卷期：v.68,no.19(October 1, 1996)
• ISSN：1520-6882

文摘

Products formed during cyanogen bromide (CNBr) digestion of -endorphin, -endorphin, and horse heart myoglobin are examined using reversed-phase high-performance liquid chromatography and electrospray massspectrometry. It is demonstrated that unstableintermediate reaction products may be formed, as well as oxidizedproducts when the CNBr reaction is performed in 0.1%TFA in water/acetonitrile (6:4 v/v) and that, under otherconditions commonly employed for the CNBr cleavagereaction, unstable intermediate products are also generated. The formation of the expected cleavage productsisfound to be improved by adjusting the hydrolysis conditions. The structure of the intermediate formed from-endorphin is examined using electrospray mass spectrometry in combination with low-energy collision-induceddissociation and tandem mass spectrometry and is shownto have a cyclic hydrated homoserine iminolactone part.The results obtained in this study explain theformationof partially cleaved proteins in the case of Met-Thr-containing sequences, which likely have a cyclic hydratedhomoserine iminolactone part instead of the putativehomoserine residue.