Thiazolinones, rearrangement

GLC is an important adjunct to protein sequence determination. Automatic "sequenators" based upon the approach developed by Edman are available and have been described in detail by Niall (60). The Edman degradation, summarized in Equation 9.5, makes use of methyl or phenylisothiocyanate which reacts with the N-terminus of a peptide. Exposure of the isothiocyanate derivative of the protein to acid results in cleavage of the terminal amino acid as a thiaxolinones and exposure of the next amine group on the peptide. Thus, the process can be repetitively carried out, each amino acid removed from the peptide, in a sequential manner. Thiazolinones rearrange in acid medium to form thiohydantoin derivatives of amino acids, some of which may be directly gas chromatographed others must be derivatized typically as trimethylsilyl derivatives. 

Then, on treatment of that derivative with trifluoroacetic acid (F3CCO2H), protonation followed by cyclization takes place and an anilinothiazoUnone is generated. On warming, the thiazolinone rearranges to the corresponding phenylthiohydantoin of alanine (Ala, A) and the peptide chain is left with a phenylalanine (Phe, F) at the terminus. 

Under the conditions of the reaction, the thiazolinone rearranges by a multistep pathway to form an A/-phenylthiohydantoin (PTH). This product contains the original N-terminal amino acid. 

The thiazolinone rearranges in dilute acid to a more stable phenylthiohydantoin (PTH)... 

Thiazolin-5-one, 2-alkoxy-4-arylazo-rearrangements, 5, 777 2-Thiazolin-5-one, 4-methyl-2-phenyl-protomeric equilibrium, 6, 249 4-Thiazolin-2-one, 4-aryl-reactions, 6, 286 4-Thiazolin-2-one, 3,4-dimethyl-protonation, 6, 286 4-Thiazolin-2-one, 4-methyl-reactions, 6, 286 Thiazolinones electrophilic attack, 5, 99 Thiazolin-2-ones IR spectroscopy, 6, 241 nucleophilic displacement, 5, 100 2-Thiazolin-4-ones reactions, 6, 287 2-substituted synthesis, 6, 306 synthesis, 5, 129 6, 309, 310 tautomerism, 6, 248 2-Thiazolin-5-ones IR spectroscopy, 6, 242 reactions, 6, 288 synthesis, 5, 138 tautomerism, 6, 249 4-Thiazolin-2-ones synthesis, 6, 314 4-Thiazolin-3-ylacetic acid esters... 

Thus, the A-terminal amino acid can be identified by analysis of the thiazolinone, and the process can be repeated on the one-unit-shortened polypeptide chain. Under the acidic conditions, the thiazolinone is actually unstable, and rearranges to a... 

The most useful method of N-terminal analysis is called the Edman degradation. This method allows the N-terminal amino acid to be removed and its identity to be determined without hydrolyzing the other peptide bonds. The reaction initially produces a thiazolinone, which is rearranged by aqueous acid to a phenylthiohydantoin for identification by high-performance liquid chromatography ... 

There are several ways to identify the N-terminal amino acid of a peptide or protein. One of the most widely used methods is to treat the protein with phenyl isothiocyanate (PITC), more commonly known as Edman s reagent. This reagent reacts with the N-terminal amino group, and the resulting thiazolinone derivative is cleaved from the protein under mildly acidic conditions. The thiazolinone derivative is extracted into an organic solvent and in the presence of acid, rearranges to a more stable phenylthiohydantoin (PTH). 

Mechanism 28.2 illustrates some of the key steps of the Edman degradation. The nucleophilic N-terminal NH2 group adds to the electrophilic carbon of phenyl isothiocyanate to form an A-phenylthiourea, the product of nucleophilic addition (Part [1]). Intramolecular cyclization followed by elimination results in cleavage of the terminal amide bond in Part [2] to form a new peptide with one fewer amino acid. A sulfur heterocycle, called a thiazolinone, is also formed, which rearranges by a multistep pathway (Part [3]) to form an A-phenylthiohydantoin. The R group in this product identifies the amino acid located at the N-terminal end. 

As shown in Mechanism 28.2, the final steps in the Edman degradation result in rearrangement of a thiazolinone to an A/-phenylthiohydantoin. Draw a stepwise mechanism for this acid-catalyzed reaction. 

Rearrangement of a penicillin sulfoxide having the sulfonylurea side chain (230) produced a different spectrum of products (R. D. G. Cooper, 1970, unpublished results), namely, the thiazolinone (231) and the 2-methoxythiazoline (232). Presumably, these arose from intermediate 233 when, instead of the normal elimination of water to produce the thia-zoline, toluene sulfonamide was eliminated to yield 231. The 2-meth-oxythiazoline was then formed by methylation of 231 with trimethyl-phosphite. 

---