โ Scribed by M.C.C. Kao; M.C.M. Chung
No coin nor oath required. For personal study only.
A procedure is described for using an inert polyvinylidene difluoride type membrane (e.g., ProBlott) as a matrix for on-membrane gas-phase pyridylethylation and as a desalting (cleanup) support following the solution phase pyridylethylation of cysteine residues in proteins. As a desalting support for (200 \mathrm{pmol}) of cardiotoxin, the membrane-derived sample gave a better recovery of phenylthiohydantoin (PTH)-amino acids during (\mathrm{N})-terminal sequencing compared to the sample obtained by rpHPLC. Using on-membrane gas-phase pyridylethylation, followed by replacement of glassfiber filter disc and membrane washing/drying in vacuo, it was possible to eliminate the broad background peak that coeluted with PTH-Ala in the HPLC chromatogram completely. Using this modified procedure, there was no evidence of (\mathrm{N})-terminal alkylation of the amino-termini of proteins. 1993 Academic Press, Inc.
๐ SIMILAR VOLUMES
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 positio