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Uncoupled analysis of secondary and tertiary protein structure by circular dichroism and electrospray ionization mass spectrometry
Author(s) -
Grandori Rita,
Matecko Irena,
Müller Norbert
Publication year - 2002
Publication title -
journal of mass spectrometry
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.475
H-Index - 121
eISSN - 1096-9888
pISSN - 1076-5174
DOI - 10.1002/jms.272
Subject(s) - chemistry , circular dichroism , electrospray ionization , mass spectrometry , myoglobin , protein tertiary structure , protein secondary structure , crystallography , electrospray , cytochrome c , molten globule , analytical chemistry (journal) , chromatography , organic chemistry , biochemistry , mitochondrion
An Erratum has been published for this article in Journal of Mass Spectrometry 38 (3) 2003, 345. Electrospray ionization mass spectrometry (ESI‐MS) applied to protein conformational studies is a powerful new method that seems to provide specific information about protein tertiary structure. In this study, we analyzed the effect of trifluoroethanol (TFE) on a myoglobin peptide and cytochrome c (cyt c ) at low pH by circular dichroism (CD) and ESI‐MS. These experiments show that coil‐to‐helix transition per se does not affect ESI mass spectra, confirming that this technique is insensitive to the local conformation of the polypeptidic chain and, rather, reports on the tertiary contacts characterizing different protein conformations. This property makes ESI‐MS an excellent method, complementary to CD, for the characterization of protein conformational changes. Fluorinated alcohols have been suggested to induce molten globule formation in acid‐unfolded cyt c . The experiments described here show that TFE does not induce major changes in the ESI mass spectrum of cyt c at pH 2.2, indicating that no stabilization of compact, globular structures is detectable under the conditions employed. On the other hand, even low concentrations of TFE (2–5%) are shown to destabilize the folded state of the protein around the mid‐point of its acid‐induced unfolding transition. Copyright © 2001 John Wiley & Sons, Ltd.