Effect of ascorbic acid in dough: reaction of oxidized glutathione with reactive thiol groups of wheat glutelin.
The reactions of oxidized glutathione generated from endogenous glutathione by the addition of ascorbic acid (AA) prior to dough mixing on free thiol groups of gluten proteins have been investigated. A small amount of (35)S-labeled glutathione was added as a tracer to identify the reaction products of GSSG and free protein thiols by radioactivity measurement. First, gluten was isolated from the dough, then the gliadins were extracted, and residual glutenin was partially hydrolyzed with thermolysin. After preseparation by gel permeation chromatography, the fractions with the highest radioactivity were separated by high-performance liquid chromatography. Radioactive peptides were identified, isolated, sequenced, and assigned to amino acid sequences of gluten protein components. The isolated peptides contained exclusively the cysteine residues C(b) and C(x) of low molecular weight subunits of glutenin, which are supposed to be highly reactive in forming intermolecular disulfide bonds. From these results it can be assumed that the cysteine residues C(b) and C(x) of the low molecular weight subunits of glutenin are at least partly present in the thiol form in flour. During dough mixing they are converted to protein-protein disulfides or glutathione-protein mixed disulfides by thiol/disulfide interchange reactions. Oxidized glutathione necessary for this reaction is generated from glutathione by the action of AA. These results are in accordance with the major hypothesis about the mechanism of action of AA.
Koehler
Peter
P
German Research Center for Food Chemistry, Lichtenbergstrasse 4, D-85748 Garching, Germany. peter.koehler@Lrz.tum.de.
D000595:Amino Acid Sequence; D001205:Ascorbic Acid / Q000032:analysis / Q000737:chemistry*; D001939:Bread / Q000032:analysis*; D002851:Chromatography, High Pressure Liquid; D005978:Glutathione / Q000737:chemistry*; D005983:Glutens / Q000031:analogs & derivatives* / Q000737:chemistry* / Q000302:isolation & purification / Q000378:metabolism; D006868:Hydrolysis; D008969:Molecular Sequence Data; D010084:Oxidation-Reduction; D013438:Sulfhydryl Compounds / Q000737:chemistry*; D014908:Triticum / Q000737:chemistry*
D013438:Sulfhydryl Compounds; D005983:Glutens; C037302:glutenin; D005978:Glutathione; D001205:Ascorbic Acid