Peptides chemical cleavage

The most commonly utilized chemical cleavage agent is cyanogen bromide (it cleaves the peptide bond on the carboxyl side of methionine residues). V8 protease, produced by certain staphylococci, along with trypsin are two of the more commonly used proteolytic-based fragmentation agents. 

Since then, catalytic antibodies which catalyze different chemical reactions have been described. The reactions range from ester or carbonate hydrolysis to carbon-carbon bond forming reactions, bimolecular amide formation or peptide bond cleavage, so the application of catalytic antibodies to general synthetic organic chemistry seems to be very promising [22]. 

Some of the multiple-cystine peptides are resistant to enzymatic and/or chemical cleavage, and thus the more simple methods described in Section 6.1.6.2 cannot be applied. In these cases assignment of their disulfide connectivities is generally attempted by NMR structural analysis. Thereby, if the two half-cystine residues are located on opposite sides of the NMR-derived 3D structure, a disulfide bond between them is improbable. 

Chemical cleavage at selective amino acid residues is an alternative method for scission of peptide bonds in target molecules. Cleavage at the Met-Xaa bond is achieved by CNBr, at the Trp-Xaa bond by the tryptophan-directed reagent 3-bromo-3-methyl-2-[(2-nitro-phenyl)sulfanyl]-3//-indole (BNPS-skatole) and at the Asp-Xaa bond by 2% formic acid.123 24 All these reactions are carried out at acidic pH under standard conditions for each reagent. 

Breaking Disulfide Bonds Disulfide bonds interfere with the sequencing procedure. A cystine residue (Fig. 3-7) that has one of its peptide bonds cleaved by the Edman procedure may remain attached to another polypeptide strand via its disulfide bond. Disulfide bonds also interfere with the enzymatic or chemical cleavage of the polypeptide. Two approaches to irreversible breakage of disulfide bonds are outlined in Figure 3-26. 

Sonderegger, P., Jaussi, R, Gehring, H, Brunschweiler, K, and Christen, P (1982) Peptide mapping of protein bands from polyacrylamide gel electrophoresis by chemical cleavage in gel pieces and re-electrophoresis Anal Biochem 122,298—301... 

Arginine residues are very resistant to most of the usual reagents used for chemical modification. King (166) has reported the conversion of these residues to S-N- (2-pyrimidinyl) ornithine by treatment with malonaldehyde in 10 N HC1. Essentially complete modification of the 4 arginine residues in RNase was obtained. Peptide bond cleavage and disulfide interchange also occurred, however, and no conclusions are warranted on the relation of the properties of the product to the actual arginine modification. 

Since the fusion protein partner is substantially larger than the size of the required peptide, overall peptide yield represents only a fraction of the purified fusion protein, even before losses due to subsequent purification. For example, for a fusion protein of 150 amino acids and a peptide of 15 amino acids, the relative levels of products are 91 and 9%, respectively. So, it is useful to increase the proportion of the final fusion protein that comprises peptide sequences. This can be achieved either by using a smaller fusion protein or by increasing the number of peptide sequences cloned in tandem with the fusion partner. As shown in the following examples both approaches have been adopted, but with variable results. Finally, the separation of the tandem repeats of peptides into monomers can be achieved by either chemical cleavage or enzymatic cleavage (Section 1.1.1.3). 

Azari and Feeney (7) suggested that human serum transferrin and chicken ovotransferrin underwent structural changes on chelation of metal ions which stabilized the molecule to denaturation and proteolysis. This later was extended (8) to include stabilization to chemical cleavage or modification. A similar interpretation also was made by Glazer and McKenzie (55) who further suggested that the iron complex might provide two crosslinks between widely separated sections of the peptide chains. 

Since in peptides and proteins alkyl halides at pH 2.8 react only with the sulfur of methionine (Gundlach et al, 1959b), this procedure permits specific chemical cleavage of methionyl peptide bonds. 

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