Hydroxyl radical probe of protein surfaces using synchrotron X-ray radiolysis and mass spectrometry.

Abstract:

:A new approach is reported that combines synchrotron radiolysis and mass spectrometry to probe the structure of proteins. Hydroxyl radicals produced upon the radiolysis of protein solutions using synchrotron light modify amino acid side-chains on millisecond timescales. This results in the formation of stable oxidation products where the level of oxidation at the reactive residues is influenced by the accessibility of their side-chains to the bulk solvent. The aromatic and sulphur-containing residues have been found to react preferentially in accord with previous peptide studies. The sites of oxidation have been determined by tandem mass spectrometry. The rate of oxidation at these reactive markers has been measured for a number of proteolytic peptides as a function of exposure time based on the relative proportion of modified and unmodified peptide ions detected by mass spectrometry. Oxidation rates correlate closely with a theoretical measure of the accessibility of residue side-chains to the solvent in the native protein structure. This approach can distinguish the relative accessibility of the tryptophan residue side-chains of lysozyme at positions 62 and 123 from each other and all other tryptophan residues, and phenylalanine at position 34 from phenylalanine residues at positions 3 and 38 based upon their rates of oxidation.

journal_name

Int J Radiat Biol

authors

Kiselar JG,Maleknia SD,Sullivan M,Downard KM,Chance MR

doi

10.1080/09553000110094805

subject

Has Abstract

pub_date

2002-02-01 00:00:00

pages

101-14

issue

2

eissn

0955-3002

issn

1362-3095

journal_volume

78

pub_type

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