Oxidation of methionine residues in polypeptide ions via gas-phase ion/ion chemistry

Abstract

The gas-phase oxidation of methionine residues is demonstrated here using ion/ion reactions with periodate anions. Periodate anions are observed to attach in varying degrees to all polypeptide ions irrespective of amino acid composition. Direct proton transfer yielding a charge-reduced peptide ion is also observed. In the case of methionine and, to a much lesser degree, tryptophan-containing peptide ions, collisional activation of the complex ion generated by periodate attachment yields an oxidized peptide product (i.e., [M + H + O](+)), in addition to periodic acid detachment. Detachment of periodic acid takes place exclusively for peptides that do not contain either a methionine or tryptophan side chain. In the case of methionine-containing peptides, the [M + H + O](+) product is observed at a much greater abundance than the proton transfer product (viz., [M + H](+)). Collisional activation of oxidized Met-containing peptides yields a signature loss of 64 Da from the precursor and/or product ions. This unique loss corresponds to the ejection of methanesulfenic acid from the oxidized methionine side chain and is commonly used in solution-phase proteomics studies to determine the presence of oxidized methionine residues. The present work shows that periodate anions can be used to 'label' methionine residues in polypeptides in the gas phase. The selectivity of the periodate anion for the methionine side chain suggests several applications including identification and location of methionine residues in sequencing applications.

Figure 1

Spectra illustrating gas-phase covalent modification of ARAMAKA, including (a) ion/ion reaction between doubly protonated peptide cation and periodate anion, (b) CID of the isolated ion/ion complex producing the [M+H+O]⁺ species, (c) MS³ of the oxidized peptide, and (d) MS⁴ of the [b₆+O-64]⁺ identifying the site of modification as the methionine residue. Degree symbols (°) denote water losses whereas asterisks (*) denote ammonia losses. Squares (□) denote fragments that have lost the modified methionine side chain, e.g., y ^□₈ corresponds to [y₈+O-HSOCH₃]⁺. The lightning bolt () is used to denote the ion that has been subjected to CID.

Figure 2

Spectra illustrating activation of ion/ion complexes produced via reactions between periodate anions and doubly protonated (a) KGAILMGAILR, (b) substance P, (c) MHRQETVDC, and (d) an eleven-residue segment of β-amyloid peptide.

Figure 3

Ion trap CID of the [M+H+O]+ species for reactions between periodate anion and doubly protonated (a) KGAILMGAILR, (b) substance P, (c) MHRQETVDC, and (d) β-amyloid peptide residues 25-35. Degree symbols (°) denote water losses whereas asterisks (*) denote ammonia losses. The lightning bolt () is used to denote the ion that has been subjected to CID.

Figure 4

Spectra illustrating activation of ion/ion complexes produced via reactions between periodate anions and doubly protonated (a) ARAAAKA, (b) ARACAKA, and (c) angiotensin II.

Figure 5

Spectra illustrating activation of ion/ion complexes produced via reactions between periodate anions and doubly protonated (a) ARAWAKA, (b) Trp-11 Neurotensin, and (c) GLSDGEWQQVLNVWGK.

Figure 6

Comparison of solution-phase and gas-phase oxidation of the peptide ARAMAKA. (a) CID of the [M+H+O]⁺ species produced via solution-phase reaction with sodium periodate, and (b) MS³ of the [M+H+O]⁺ from dissociation of the [M+2H+IO₄^-]⁺ species produced by the gas-phase ion/ion reaction of doubly protonated ARAMAKA with periodate anion.

Figure 7

a) Ion/ion reaction between doubly protonated GRGMGRGMGRL and periodate anions. Product ion spectra derived from b) ion trap CID of [M+3H+2IO₄^-]⁺, c) further activation of [M+H+2(O)]⁺, and d) activation of the 64 Da loss from the doubly oxidized species,[M+H+2(O)-HSOCH₃]⁺. Degree symbols (°) denote water losses whereas asterisks (*) denote ammonia losses. Open squares (□) denote fragments that have lost one modified methionine side chain while closed squares (■) denote fragments that have lost two modified methionine side chains, e.g., y₈^□ corresponds to [y₈ +O-HSOCH ⁺+O-HSOCH₃]⁺ y₈^■ corresponds to [y₈+O-2HSOCH₃]⁺. The lightning bolt () is used to denote the ion that has been subjected to CID.

Scheme 1

Mechanism of rearrangement of the oxidized methionine side chain to produce loss of methanesulfenic acid (64 Da).[18-21]

Scheme 2

Proposed mechanism for ion/ion reaction between periodate anion and a doubly cationic methionine-containing peptide to form the oxidized species. Adapted from references 42 and 43.