Reduction of disulfide bonds in proteins by 2-aminothiophenol under weakly acidic conditions.
We developed a method for reducing disulfide bonds in proteins under weakly acidic conditions by use of 2-aminothiophenol. The disulfide bonds in hen egg-white lysozyme, ribonuclease A, and soybean trypsin inhibitor were quantitatively reduced by 2-aminothiophenol in phosphate buffer, pH6, containing 8 M Gdn HCl, 1 mM EDTA, and 20% ethanol, for 60 min at 40 degrees C. On analysis of the RP-HPLC patterns of tryptic peptides, which were derived from reduced and S-alkylated lysozyme and ribonuclease A at pH 6, it was confirmed that no side reaction occurred. Moreover, the reduction under weakly acidic conditions was demonstrated to be applicable for the location of such a labile residue as O-acetylated tyrosine.
Abe
Y
Y
Faculty of Pharmaceutical Sciences, Kyushu University, Fukuoka.
Ueda
T
T
Imoto
T
T
D000814:Aniline Compounds / Q000737:chemistry*; D000818:Animals; D002645:Chickens; D003545:Cysteine / Q000031:analogs & derivatives / Q000737:chemistry; D004220:Disulfides / Q000737:chemistry*; D006863:Hydrogen-Ion Concentration; D009113:Muramidase / Q000737:chemistry; D010084:Oxidation-Reduction; D011506:Proteins / Q000737:chemistry*; D013388:Succinimides / Q000737:chemistry; D014357:Trypsin
D000814:Aniline Compounds; D004220:Disulfides; D011506:Proteins; C087317:S-succinimidocysteine; D013388:Succinimides; C005464:2-aminothiophenol; C017212:S-(1,2-dicarboxyethyl)cysteine; D009113:Muramidase; D014357:Trypsin; D003545:Cysteine