Simultaneous racemization and isomerization at specific aspartic acid residues in alpha B-crystallin from the aged human lens.
We provide evidence that the racemization and isomerization of aspartyl(Asp) residues occur simultaneously in the alpha B-crystallin in the lens of aged (mean age: 80 years) and young (age: 11 months) humans. We purified alpha B-crystallin and subjected it to tryptic digestion. The resulting peptides were separated by reverse-phase high-performance chromatography (RP-HPLC) and were characterized by amino-acid composition, sequence analysis and mass spectrometry. Two specific sites, Asp-36 (D/L of Asp: 0.92) and Asp-62(D/L of Asp: 0.57), among 13 Asp/asparginyl (Asn) residues in aged alpha B-crystallin, were found to be highly racemized and isomerized to form beta-Asp residues. The beta-Asp-containing peptides were clearly distinguished from normal Asp-containing (alpha-Asp) peptides by RP-HPLC. The racemization and isomerization of Asp residues in aged alpha B-crystallin may occur via a succinimide intermediate. In young alpha B-crystallin, we observed neither racemization nor isomerization. We also found that Met-68 was oxidized to form Met sulfoxide to a greater extent in aged alpha B-crystallin than in young alpha B-crystallin. We concluded that racemization, isomerization, and oxidation of alpha B-crystallin occur spontaneously in the aging process.
Fujii
N
N
Discovery Research Division, Takeda Chemical Industries, Ltd., Ibaraki, Japan.
Ishibashi
Y
Y
Satoh
K
K
Fujino
M
M
Harada
K
K
D000368:Aged; D000369:Aged, 80 and over; D000375:Aging; D000595:Amino Acid Sequence; D000596:Amino Acids / Q000032:analysis; D001224:Aspartic Acid / Q000737:chemistry*; D003459:Crystallins / Q000737:chemistry* / Q000302:isolation & purification; D006801:Humans; D007223:Infant; D007536:Isomerism; D008715:Methionine / Q000737:chemistry; D008969:Molecular Sequence Data; D010084:Oxidation-Reduction; D010455:Peptides / Q000737:chemistry; D013237:Stereoisomerism; D014357:Trypsin
D000596:Amino Acids; D003459:Crystallins; D010455:Peptides; D001224:Aspartic Acid; D008715:Methionine; D014357:Trypsin