Identification of hydrogen peroxide oxidation sites of alpha A- and alpha B-crystallins.
The alpha-crystallins are the most abundant structural proteins of the lens and, because of their chaperone activity, contribute to the solubility of the other crystallins. With aging, the lens crystallins undergo a variety of modifications which correlate with a loss of solubility and the development of cataract. A recent study demonstrating that alpha-crystallins exposed in vitro to FeCl3 and H2O2 exhibit decreased chaperone activity, implicates metal catalyzed oxidations of alpha-crystallins in this loss of solubility. The present study has determined that alpha-crystallins incubated with FeCl3 and H2O2 are modified by the nearly complete oxidation of all methionine residues to methionine sulfoxide, with no other detectable reaction products. The modifications were identified from the molecular weights of peptides formed by enzymatic digestion of the alpha-crystallins and located by tandem mass spectrometric analysis of the fragmentation pattern of the mass spectra of the fragments from peptides with oxidized methionine is loss of 64 Da, which corresponds to loss of CH3SOH from the methionine sulfoxide. These fragments are useful in identifying peptides that include oxidized methionine residues.
Smith
J B
JB
Department of Chemistry, University of Nebraska, Lincoln 68588-0304, USA. jbsmith@unlinfo.unl.edu
Jiang
X
X
Abraham
E C
EC
D000595:Amino Acid Sequence; D000818:Animals; D002417:Cattle; D002712:Chlorides; D002851:Chromatography, High Pressure Liquid; D003459:Crystallins / Q000378:metabolism*; D004591:Electrophoresis, Polyacrylamide Gel; D005290:Ferric Compounds / Q000378:metabolism; D006360:Heat-Shock Proteins / Q000378:metabolism; D006861:Hydrogen Peroxide / Q000378:metabolism*; D013058:Mass Spectrometry; D008715:Methionine / Q000031:analogs & derivatives / Q000378:metabolism; D018832:Molecular Chaperones / Q000378:metabolism*; D008969:Molecular Sequence Data; D008970:Molecular Weight; D010084:Oxidation-Reduction; D010449:Peptide Mapping
D002712:Chlorides; D003459:Crystallins; D005290:Ferric Compounds; D006360:Heat-Shock Proteins; D018832:Molecular Chaperones; D008715:Methionine; D006861:Hydrogen Peroxide; C024555:ferric chloride; C013111:methionine sulfoxide