Application of S-pyridylethylation of cysteine to the sequence analysis of proteins

✍ Scribed by A.S. Mak; B.L. Jones

• Publisher: Elsevier Science
• Year: 1978
• Tongue: English
• Weight: 488 KB
• Volume: 84
• Category: Article
• ISSN: 0003-2697
• DOI: 10.1016/0003-2697(78)90061-1

✦ Synopsis

Methods are given for determination of the phenylthiohydantoin derivative of pyridylethylcysteine (PTH-PECys) by gas chromatography, both with and without prior trimethylsilylation.

Using the high-cysteine polypeptide purothionin, obtained from wheat, as a model system, cysteine residues at positions 3, 4, 12, 16, and 25 were unambiguously determined and those at positions 29 and 31 were indicated. The PTH-PECys remains in the aqueous phase during the ethyl acetate extraction step and neither PTH-His nor PTH-Arg interferes with the PTH-PECys determination. The uv spectrum of PTH-PECys differs from the spectra of PTH-Arg and PTH-His, allowing uv spectroscopy to be used as another method for determining which Edman degradation cycles yielded PTH-PECys. The susceptibility of PECys peptide bonds to enzymatic hydrolysis was also examined. Chymotrypsin hydrolyzes purothionin at peptide bonds connecting some PECys and Arg residues. Carboxypeptidase B catalyzes the hydrolysis of PECys from the C terminus of peptides, but carboxypeptidase A does not. ' Contribution No.