Using Metal-Catalyzed Oxidation Reactions and Mass Spectrometry to Identify Amino Acid Residues Within 10 Å of the Metal in Cu-Binding Proteins

Juma D. Bridgewater, Jihyeon Lim, Richard W. Vachet

Research output: Contribution to journalArticle

26 Citations (Scopus)

Abstract

Metal-catalyzed oxidation (MCO) reactions and mass spectrometry (MS) can be used together to determine the amino acids bound to Cu in a metalloprotein. Selective oxidation of only amino acids bound to Cu during the MCO/MS approach relies on proper choice of the types and concentrations of the reducing and oxidizing agents. We show here that if these MCO reagent concentrations are "detuned" or varied slightly from optimal conditions, nonmetal-bound amino acids close to Cu can also be oxidized in a controlled manner. Using Cu/Zn superoxide dismutase as a model system, we demonstrate that amino acids up to 10 Å from Cu can be modified as long as they are readily accessible to diffusing reactive oxygen species. UV/VIS spectroscopy and measurements of oxidation kinetics provide evidence that the protein's structural integrity around Cu is maintained during the detuned MCO reactions. Because this method can identify amino acids around Cu that have low solvent accessibility, it should complement other radical-based protein surface-mapping techniques.

Original languageEnglish (US)
Pages (from-to)1552-1559
Number of pages8
JournalJournal of the American Society for Mass Spectrometry
Volume17
Issue number11
DOIs
StatePublished - Nov 2006
Externally publishedYes

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Mass spectrometry
Mass Spectrometry
Carrier Proteins
Metals
Amino Acids
Oxidation
Metalloproteins
Nonmetals
Reducing Agents
Oxidants
Structural integrity
Reactive Oxygen Species
Ultraviolet spectroscopy
Spectrum Analysis
Membrane Proteins
Kinetics
Proteins

ASJC Scopus subject areas

  • Structural Biology
  • Spectroscopy

Cite this

Using Metal-Catalyzed Oxidation Reactions and Mass Spectrometry to Identify Amino Acid Residues Within 10 Å of the Metal in Cu-Binding Proteins. / Bridgewater, Juma D.; Lim, Jihyeon; Vachet, Richard W.

In: Journal of the American Society for Mass Spectrometry, Vol. 17, No. 11, 11.2006, p. 1552-1559.

Research output: Contribution to journalArticle

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