N-terminal derivatization

Fig. 3. Automatically acquired MALDI-TOF/TOF MS/MS spectra of a tryptic peptide with m/z 1136.67 (upper panel) and of the same peptide after derivatization with SPITC at m/z 1393.11 (lower panel) using CHCA matrix. Whereas the unmodified peptide displays a rather complex spectrum with intense b- and y-ions in the same mass range, the lower spectrum shows a complete and dominant y-series allowing an easy manual sequence interpretation from m/z 189.09 to m/z 1178.32. The peptide sequence LRGGLC YLGK was deduced from the y-series with guanidated Lys (homoarginine = Flar) and N-terminal derivatization with SPITC. The cysteine residue was reduced and alkylated with DTT and iodoacetamide (M = 160.03 g/mol).

N-Terminal Derivatization with 2-Sulfobenzoic Acid Cyclic Anhydride... 

Numerous chemical derivatization strategies have been developed also to direct the fragmentation reactions of protonated peptides toward the selective formation of single characteristic sequence-type product ions [85-87]. For example, Sununerfield and coworkers have demonstrated that N-terminal derivatization, with phenyliso-thiocyanate to form the corresponding phenylthiocarbamoyl (PTC) derivative, results in exclusive fragmentation of the amide bond between the first two amino... 

We describe here on tissue N-terminal chemical derivatization of peptides performed after on tissue enzymatic digestion. Since N-terminal derivatization has proven to be more efficient than the corresponding C-terminal reactions in solution, focus has been given to N-terminal chemical derivatizations. More specifically, among the tested derivatization approaches,... 

It should be noted that the majority of the derivatization techniques modify the peptide s N-terminus. The reason is that the N-terminal amine group is easier to modify than the C-terminal carboxyl group. Also, due to differences in pKa value in the e-amino group of lysine, there are possible reaction that modify the N-terminus only, while the lysine side chains remain intact. Modifications of carboxyl groups... 

Rhodobactin (31) was isolated from Rhodococcus rhodochrous (86). A sequence of four Om units derivatized in different ways is linked together. The nitrogen atoms of the N-terminal Om are substituted with DHB groups, the N-terminal Om is followed by two Om moieties, for which the N -amino groups are transformed into urea units (NH2CONH-), and the C-terminus is cOHOm. The stereochemistry of the Orn units was not determined. Rhodobactin forms a 1 1 Fe /Lig complex. Iron uptake was studied with Fe ". ... 

Human parathyroid hormone, a 34-residue peptide, has an N-terminal Ser suitable for conversion into the /V -glyoxyl moiety, thus, permitting a subsequent thiazolidine conjugation with 1-amino-2-thiol derivatized biotin.[l7 ... 

Thiazoles are less electron-rich isosteres of pyridines and therefore it was speculated that compounds with such substitution may have improved metabolic stability [30]. The modeling of A-82200 in which the N-terminal pyridinyl group was substituted by a 4-thiazolyl moiety indicated that the 5-membered ring binds in the S3 subsite and can be further derivatized at the 2 position by an isopropyl group. The isopropyl functionality makes van der Waals contacts with Val82 and fills the hydrophobic part of the S3 subsite in nearly optimal fashion. 

Dansyl chloride has been widely utilized in peptide analyses to determine the N-terminal amino acid (60,86). Mendez et al. (87) suggested that derivatization of the peptides with dansyl... 

The reaction of 2,4-dinitrofluorobenzene (DNFB) (Sanger s reagent [10]) with amino acids is another useful technique which is often employed for the analysis of N-terminal amino acids by TLC and column chromatography after derivatization. The reaction involved in product formation is shown in Fig.4.6. The separated derivatives are determined by measuring the quenching of fluorescence on TLC plates or by UV analysis after column chromatography. The generalized absorption curves of dinitrophenyl (DNP)-amino acids in acidic and alkaline solutions are shown in Fig. 4.7. 

Phenyl isocyanate < OVn=c=o Used for derivatization of N-terminal peptide residue... 

N-Terminal Peptide Derivatization Using 2-Sulfobenzoic Acid... 

That the N-terminal methionine is exposed to the solvent was also concluded from studies directed toward the reduction and alkylation of the disulfides of rhGH. The conditions that were found to be optimal for the derivatization of the four sulfhydryls of rhGH resulted in the production of a side reaction when the procedure was applied to the methionyl analog. This side product was identified as containing carboxymethyl-S-methionine at the amino terminal residue. 

Derivatizations in the N-terminal sequence of human angiotensinogen led to weakly active renin inhibitors 183). According to Szelke et al.l84), highly active substrate analogs with modified peptide linkage ( CH2 NH—) were formed by reduction of the CO—NH bond in Leu-Leu or Phe-Phe (Table 2). 

N-terminal end) The end of the peptide chain with a free or derivatized amino group. As the peptide is written, the N terminus is usually on the left. The carboxyl group of the N-terminal amino acid links it to the rest of the peptide, (p. 1174)... 

A method for determining the N-terminal amino acid of a peptide. The peptide is treated with 2,4-dinitrofluorobenzene (Sanger s reagent), then completely hydrolyzed. The derivatized amino acid is easily identified, but the rest of the peptide is destroyed in the hydrolysis, (p. 1180)... 

Interpretation of the fragmentation spectra can be difficult and time consuming. It can be simplified if the peptide is modified in such a way that one type of fragmentation is favoured. This can be achieved by derivatizing the amino terminal group with N-succinimidyl-2-(3-pyridyl) acetate, which promotes b fragments [59], Modification of the peptide also can make the distinction between C- and N-terminal fragments easier. 

Derivatization also allows specific residues to be detected and located by comparing the spectra before and after derivatization. The derivatizations most often used for this purpose are the formation of ethyl esters from Asp, Glu and C-terminal carboxylic acids acetylation of the N-terminal amino group and Lys and the Edman reaction for the N-terminal amino acid [60],... 

---