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Native Mass Spectrometry of Iron-Sulfur Proteins

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Fe-S Proteins

Part of the book series: Methods in Molecular Biology ((MIMB,volume 2353))

Abstract

Iron-sulfur clusters constitute a large and widely distributed group of protein cofactors that play key roles in a wide range of metabolic processes. The inherent reactivity of iron-sulfur clusters toward small molecules, for example, O2, NO, or free Fe, makes them ideal for sensing changes in the cellular environment. Nondenaturing, or native, MS is unique in its ability to preserve the noncovalent interactions of many (if not all) species, including stable intermediates, while providing accurate mass measurements in both thermodynamic and kinetic experimental regimes. Here, we provide practical guidance for the study of iron-sulfur proteins by native MS, illustrated by examples where it has been used to unambiguously determine the type of cluster coordinated to the protein framework. We also describe the use of time-resolved native MS to follow the kinetics of cluster conversion, allowing the elucidation of the precise series of molecular events for all species involved. Finally, we provide advice on a unique approach to a typical thermodynamic titration, uncovering early, quasi-stable, intermediates in the reaction of a cluster with nitric oxide, resulting in cluster nitrosylation.

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Acknowledgments

We thank the University of East Anglia for funding the purchase of the electrospray ionization mass spectrometer instrument. This work has been supported by a series of grants from the BBSRC over several years, most recently BB/P006140/1 and BB/P006140/1.

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  1. Centre for Molecular and Structural Biochemistry, School of Chemistry, University of East Anglia, Norwich, UK

    Jason C. Crack & Nick E. Le Brun

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  1. Jason C. Crack
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  2. Nick E. Le Brun
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Correspondence to Nick E. Le Brun .

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  1. Department of Chemistry, Wake Forest University, Winston Salem, NC, USA

    Patricia C. Dos Santos

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Crack, J.C., Le Brun, N.E. (2021). Native Mass Spectrometry of Iron-Sulfur Proteins. In: Dos Santos, P.C. (eds) Fe-S Proteins. Methods in Molecular Biology, vol 2353. Humana, New York, NY. https://doi.org/10.1007/978-1-0716-1605-5_13

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  • DOI: https://doi.org/10.1007/978-1-0716-1605-5_13

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  • Publisher Name: Humana, New York, NY

  • Print ISBN: 978-1-0716-1604-8

  • Online ISBN: 978-1-0716-1605-5

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