Phenylthiohydantoin

Phenylthiohydantoin (PTH) refers to a class of derivatives formed during the Edman degradation method used for sequencing peptides and proteins. In this process, the N-terminal amino acid reacts with phenylisothiocyanate to form a phenylthiocarbamoyl (PTC) intermediate. Under mildly basic conditions this intermediate cyclizes and, upon acid treatment, is cleaved to yield the phenylthiohydantoin derivative of the N-terminal amino acid. The resulting PTH-amino acid is then isolated and identified, typically by chromatographic methods.

The PTH derivatives are characterized by a hydantoin ring bearing a phenylthio substituent. Each standard amino acid produces a distinct PTH derivative, allowing identification of the removed N-terminal residue in successive Edman cycles. The separation and identification are commonly performed by high-performance liquid chromatography (HPLC) with ultraviolet detection due to the aromatic phenyl group, and increasingly by mass spectrometry.

Applications and scope include procedural sequencing of peptides and proteins, particularly in older or specialized workflows where partial sequence information is sufficient or where intact N-termini are preserved. PTH derivatives are central to the original Edman degradation technique, which removes one residue per cycle to build a sequence from the N-terminus.

Limitations and practical notes: Edman degradation with PTH derivatives is most effective for relatively short peptides and requires an unblocked N-terminus. Certain residues, such as proline at the N-terminus, can hinder the reaction, and the process becomes less practical for long sequences. Safety considerations apply to reagents such as phenylisothiocyanate.

History: The method and the use of PTH derivatives were developed by Marcel Edman in the 1950s and established a foundational approach to protein sequencing.