PTC-peptide

The formation of the phenylthiohydantoin does not require the presence of water, as does the hydrolysis of peptide bonds. Thus by heating the PTC-peptide with anhydrous HCl in nitromethane it is possible to break off the iV-terminal residue as a phenylthiohydantoin without splitting other bonds in the peptide. The hydantoin dissolves in the nitromethane and is then separated and hydrolyzed to the amino acid which may be identified by paper chromatography. The rest of the peptide with the J T-terminal residue removed is insoluble in nitromethane and the process may be repeated. The second residue is thus split off and identified. This method has given excellent results with synthetic peptides and it will be interesting to see how far it may be applied to a... 

The sodium salt of PTC-peptide is redissolved in water (2-10 ml), and aliquots corresponding to 0.2-1,0 pM are made 3N with respect to hydrochloric acid and 0.2-1.0 x 10 M with respect to peptide by addition of the correct amounts of water and 5.7N HCl. The rate of release of phenylthi-ohydantoin can be determined by following the change of the absorption maximum of the solution (from 240 nm or lower to 265-270 nm) during a period of about 2 h. If the transformation takes place too slowly for a given peptide, the effect of increasing temperature to 40-45 C should be tried. 

A major advance was devised by Pehr Edman (University of Lund Sweden) that has become the standard method for N terminal residue analysis The Edman degrada tion IS based on the chemistry shown m Figure 27 12 A peptide reacts with phenyl iso thiocyanate to give a phenylthwcarbamoyl (PTC) denvative as shown m the first step This PTC derivative is then treated with an acid m an anhydrous medium (Edman used mtromethane saturated with hydrogen chloride) to cleave the amide bond between the N terminal ammo acid and the remainder of the peptide No other peptide bonds are cleaved m this step as amide bond hydrolysis requires water When the PTC derivative IS treated with acid m an anhydrous medium the sulfur atom of the C=S unit acts as... 

Step 1 A peptide is treated with phenyl isothiocyanate to give a phenylthiocarbamoyl (PTC) derivative... 

Step 2 On reaction with hydrogen chloride m an anhydrous solvent the thiocarbonyl sulfur of the PTC derivative attacks the carbonyl carbon of the N terminal ammo acid The N terminal ammo acid is cleaved as a thiazolone derivative from the remainder of the peptide... 

Table 8. Thermodynamic parameters of the coil-to-helix transition of collagen-model peptides, covalently linked with 1,2,3-propanetricarboxylic acid (PTC) and Lys-Lys, respectively. Solvent 1% aqueous acetic arid (pH 3.0)...

HPLC High Performance Liquid Chromatography LHRH Lutenizing Hormone Releasing Hormone PTC Phase transfer catalyst SPPS Solid Phase Peptide Synthesis TFA Trifluoro acetic acid... 

PITC has been used extensively in the sequencing of peptides and proteins and reactions under alkaline conditions with both primary and secondary amino acids. The methods of sample preparation and derivatization follow a stringent procedure which involves many labour-intensive stages. However, the resulting phenylthio-carbamyl-amino acids (PTC-AA s) are very stable, and the timing of the derivatization step is not as critical as when using OPA. 

Reaction with phenylisothiocyanate (PITC) in alkaline conditions produces stable phenylthiocarbamyl (PTC) adducts which can be detected either in the ultraviolet below 250 nm or electrochemically. However, this method involves a complex derivatization procedure and offers poorer sensitivity than the alternatives available for individual amino acids. It is useful, however, in conjunction with the automated analysis of peptides when single derivatized residues can be cleaved and analysed after conversion in acidic conditions to phenylthiohydantoins. 

To sequence an entire polypeptide, a chemical method devised by Pehr Edman is usually employed. The Edman degradation procedure labels and removes only the amino-terminal residue from a peptide, leaving all other peptide bonds intact (Fig. 3-25b). The peptide is reacted with phenylisothiocyanate under mildly alkaline conditions, which converts the amino-terminal amino acid to a phenylthiocarbamoyl (PTC) adduct. The peptide bond next to the PTC adduct is then cleaved in a step carried out in anhydrous trifluo-roacetic acid, with removal of the amino-terminal amino acid as an anilinothiazolinone derivative. The deriva-tized amino acid is extracted with organic solvents, converted to the more stable phenylthiohydantoin derivative by treatment with aqueous acid, and then identified. The use of sequential reactions carried out under first basic and then acidic conditions provides control over... 

Fig. 4.1 Overview of ribosome. Center These space filled representations are based on docking of the large ribosomal subunit [1] (grey RNA and blue protein) onto the small subunit [2] (yellow RNA and green protein) based on a protein alpha carbon and rRNA phosphate backbone trace of the 70S ribosome bound with tRNA [53]. Functional sites are circled and labeled according to the corresponding bound tRNA molecules A-site (red), P-site (orange), E-site (purple) and factor binding site (FBS). Peptide bond formation is catalyzed at the peptidyl transferase center (PTC) that is located on the large subunit between the A-site and P-site. Left The...

In order to determine the carboxy-terminus of Hez-PBAN,. 200 pmol of purified peptide was digested with carboxypeptidase P. Released amino acids were periodically analyzed as their PTC derivatives. Leu was found to be the C-terminal residue followed by Arg, Pro, Ser, Phe, Tyr, and Lys, respectively, thus confirming the automated Edman degradation data (Table I, run 5). However, none of the data could distinguish between a C-terminal amide or free acid. 

Figures 2 and 3). Peptide bond formation takes place at the peptidyl transferase center (PTC) and as the nascent peptide is extended, the single-stranded protein is extruded out the exit tunnel to further fold. One of the first... 

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