Quantification of structural alterations of L-Asp and L-Asn residues in peptides related to neuronal diseases by reversed-phase high-performance liquid chromatography.
A method for analyzing the structural alterations in Asn or Asp residues was developed by using the peptides related to neuronal conformational diseases, i.e., the prion protein (PrP)(106-126) and the Alzheimer's amyloid beta (A beta) protein(6-28). The alterations were analyzed by reversed-phase (RP) HPLC, because the peptides containing the structurally altered residues were diastereoisomers of each other, and they were separated with the mobile phase containing an MeCN/sodium phosphate solution and NaCl. The amount of L-Asp, L-isoAsp, D-Asp, or D-isoAsp residues in each PrP peptide isomer was simultaneously quantified by carrying out single HPLC analysis; these residues were generated by the deamidation of the Asn residue. Only 0.3% of the newly generated peptide containing the D-Asp residue was detected. Furthermore, the investigation of the partial fragment of the A beta protein revealed that the present method possessed the ability of simultaneous analysis of the isomerizations of two Asp residues. These results implied that the present method was highly sensitive and reduced the time required for the analysis. This method may accelerate the elucidation of the PrP and A beta protein functions, because the structural alterations of Asn and Asp have been reported to influence these functions.
Sadakane
Yutaka
Y
School of Pharmaceutical Science, Kyushu University of Health and Welfare, Nobeoka 882-8508, Japan. sadapon@phoenix.ac.jp
Konoha
Keiko
K
Kawahara
Masahiro
M
Nakagomi
Kazuya
K
D000595:Amino Acid Sequence; D016229:Amyloid beta-Peptides / Q000737:chemistry*; D001216:Asparagine / Q000032:analysis* / Q000737:chemistry; D001224:Aspartic Acid / Q000032:analysis* / Q000737:chemistry; D002493:Central Nervous System Diseases / Q000378:metabolism; D002851:Chromatography, High Pressure Liquid / Q000379:methods*; D056148:Chromatography, Reverse-Phase / Q000379:methods*; D008969:Molecular Sequence Data; D010446:Peptide Fragments / Q000737:chemistry*; D010455:Peptides / Q000737:chemistry*; D011328:Prions / Q000737:chemistry*; D013237:Stereoisomerism; D013696:Temperature
D016229:Amyloid beta-Peptides; D010446:Peptide Fragments; D010455:Peptides; D011328:Prions; C082632:prion protein (106-126); D001224:Aspartic Acid; D001216:Asparagine