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Investigations of the Mechanism of the “Proline Effect” in Tandem Mass Spectrometry Experiments: The “Pipecolic Acid Effect”

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Journal of The American Society for Mass Spectrometry

Abstract

The fragmentation behavior of a set of model peptides containing proline, its four-membered ring analog azetidine-2-carboxylic acid (Aze), its six-membered ring analog pipecolic acid (Pip), an acyclic secondary amine residue N-methyl-alanine (NMeA), and the D stereoisomers of Pro and Pip has been determined using collision-induced dissociation in ESI-tandem mass spectrometers. Experimental results for AAXAA, AVXLG, AAAXA, AGXGA, and AXPAA peptides are presented, where X represents Pro, Aze, Pip, or NMeA. Aze- and Pro-containing peptides fragment according to the well-established “proline effect” through selective cleavage of the amide bond N-terminal to the Aze/Pro residue to give yn + ions. In contrast, Pip- and NMA-fragment through a different mechanism, the “pipecolic acid effect,” selectively at the amide bond C-terminal to the Pip/NMA residue to give bn + ions. Calculations of the relative basicities of various sites in model peptide molecules containing Aze, Pro, Pip, or NMeA indicate that whereas the “proline effect’ can in part be rationalized by the increased basicity of the prolyl-amide site, the “pipecolic acid effect” cannot be justified through the basicity of the residue. Rather, the increased flexibility of the Pip and NMeA residues allow for conformations of the peptide for which transfer of the mobile proton to the amide site C-terminal to the Pip/NMeA becomes energetically favorable. This argument is supported by the differing results obtained for AAPAA versus AA(D-Pro)AA, a result that can best be explained by steric effects. Fragmentation of pentapeptides containing both Pro and Pip indicate that the “pipecolic acid effect” is stronger than the “proline effect.”

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Acknowledgments

This work was supported by the National Science Foundation, J.C.P.: (CAREER:0348889 and CHE:0911244) and the National Institutes of Health, V.H.W. (GM R0151387). Additional support was contributed by the Camille and Henry Dreyfus Foundation through the Henry Dreyfus Teacher-Scholar Award (J.C.P.) and the College of William and Mary. The authors thank Marriah Binek and Katie Henke for contributions to the project.

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  1. Omar M. Hamdy

    Present address: Department of Chemistry, University of California, Riverside, CA, 92521, USA

Authors and Affiliations

  1. Department of Chemistry, The College of William and Mary, Williamsburg, VA, 23187, USA

    Mary Disa M. Raulfs, Matthew Bernier, Omar M. Hamdy, Ashley Janiga & John C. Poutsma

  2. Department of Chemistry, University of Arizona, Tucson, AZ, 85721, USA

    Linda Breci, Matthew Bernier & Vicki Wysocki

  3. Department of Chemistry, The Ohio State University, Columbus, OH, 43210, USA

    Matthew Bernier & Vicki Wysocki

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  1. Mary Disa M. Raulfs
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Correspondence to John C. Poutsma.

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Raulfs, M.D.M., Breci, L., Bernier, M. et al. Investigations of the Mechanism of the “Proline Effect” in Tandem Mass Spectrometry Experiments: The “Pipecolic Acid Effect”. J. Am. Soc. Mass Spectrom. 25, 1705–1715 (2014). https://doi.org/10.1007/s13361-014-0953-5

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