Transpeptidation in sequence analysis. Investigations concerning a native and a synthetic hexapeptide

On sequencing a hemoglobin a peptic hexapeptide with the amino acid composition Asp2, Gly, Val, Lys2 was isolated. Cleavage of this peptide with Tos-Phe-CH2Cl-trypsin resulted in the fragments Asp-Lys, Gly-Asp, Val-Lys, Asp--Lys-Gly-Asp, Asp-Lys-Val-Lys and Val-Lys-Gly--Asp. From these data two possible but inconsistent sequences for the hexapeptide can be derived: Asp-Lys-Val-Lys-Gly-Asp and Val-Lys-Asp-Lys-Gly-Asp. Fragments obtained by other cleavage procedures, direct sequencing of the globin peptide-chain with a sequenator as well as X-ray data of this hemoglobin show, that only the sequence starting with Asp occurs in the intact protein. Therefore the peptide Asp-Lys-Gly-Asp must have been formed by transpeptidation during sequence work. In order to verify this, Asp-Lys-Val-Lys-Gly-Asp was synthesized by an unequivocal, conventional procedure. Tryptic digestion of this hexapeptide also resulted in ASP-Lys-Gly-Asp in addition to the expected fragments. Thus it has been shown for the first time, that sequencing conditions may alter the constitution of a peptide.