High-performance liquid chromatography of phenylthiocarbamyl-amino acids. Application to carboxyl-terminal sequencing of proteins.
A carboxyl-terminus microsequencing method has been developed on the basis of carboxypeptidase A digestion and derivatization of released amino acids with phenylisocyanate followed by subsequent high-performance liquid chromatographic analysis of the resulting phenylthiocarbamyl amino acids. This technique has been used to confirm the carboxyl-terminal sequence of bovine brain-derived acidic fibroblast growth factor. The method is of general application for the microsequencing of the carboxyl-termini of proteins. Since identification of derivatized amino acids was performed on the entire reaction mixture following digestion, a new chromatographic identification procedure for phenylthiocarbamyl amino acids was developed in order to resolve them from spurious peaks. This procedure can be applicable to amino acid analysis.
Department of Biochemistry and Molecular Biology, Merck Institute for Therapeutic Research, Merck Sharp and Dohme Research Laboratories, Rahway, NJ 07065.
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Gimenez-Gallego
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Thomas
D000596:Amino Acids / Q000032:analysis*; D002268:Carboxypeptidases; D002851:Chromatography, High Pressure Liquid; D006868:Hydrolysis; D010670:Phenylthiourea / Q000032:analysis*; D011506:Proteins / Q000032:analysis*; D014357:Trypsin
D000596:Amino Acids; D011506:Proteins; D010670:Phenylthiourea; D002268:Carboxypeptidases; D014357:Trypsin