Peptide oxidative modification

Oxidation modifications such as carbonylation, thiol oxidation, and aromatic hydroxylation, and Maillard glycation (the reaction of sugars with amino acid side chains) are the protein modifications most frequently reported in foodstuffs that have been subjected to thermal processing. However, condensations and eliminations of side chains or peptide backbone breakdown have also been described (95). 

The oxidative modification of peptides is a most interesting topic, but there is no suitable method available. The mthenium-catalyzed oxidation with peracetic acid provides a useful method for modification. For example, the reaction of N,C-pro-tected peptides containing glycine residues with peracetic acid in the presence of RUCI3 catalyst gives a-ketoamides 69 derived from oxidation at the Cf position of the glycine residue selectively (81%, conv. 70%) (Eq. 3.81) [139]. 

The combination NaBrOa-HBr in a ratio that produces 1 mole of bromine per mole of tryptophan peptide may prove to be useful for studies where only oxidative modification but not cleavage of tryptophan bound in a protein is required. 

The oxidation of trypsin and trypsinogen was carried out in aqueous 0.1 M acetate buffer solutions at room temperature. In this particular case and under these conditions no significant cleavage of peptide bonds next to tryptophan residues occurred. Careful analysis of hydrolyzates of NBS-oxidized trypsinogen and trypsin confirmed the selectivity of the oxidative modification of the protein, as Table XXIV shows. There is no significant loss of tyrosine, histidine, serine, threonine, or cystine, although all of these amino acids will react with NBS but considerably less rapidly than tryptophan. 

These derivatizations are highly selective, and may thus allow PSD measurements to be carried out on peptides after modification. Such a protocol would significantly enhance our ability to derive sequence information from PSD spectra, because the mass shifts observed in fragments help locate the particular residue within the peptide, and also confirm assignments of fragments as arising from N- or C-terminal regions. In addition to derivatizations that may modify the C- and N-termini and the derivatization of tyrosine residues, we have carried out oxidation of methionine residues with sufficient specificity to enable measurement of PSD spectra. 

The ammo acid sequences of five plasma kallikrem inhibitors (17-mers) are shown. The sequences of the synthetic peptides derive from the clones PK2, PK4, PK6 and PK13 (isolated in phage selections using library 1) and from clone PK15 (an affinity-matured clone isolated from library 2). Indicated are the molecular masses and the inhibitory activities before and after the modification of the peptides with tBMB. The reduced linear peptides were incubated with plasma kallikrem, and the inhibitory activity was measured immediately to minimize the risk of peptide oxidation. 

The activity of PK and NRPSs is often precluded and/or followed by actions upon the natural products by modifying enzymes. There exists a first level of diversity in which the monomers for respective synthases must be created. For instance, in the case of many NRPs, noncanonical amino acids must be biosynthesized by a series of enzymes found within the biosynthetic gene cluster in order for the peptides to be available for elongation by the NRPS. A second level of molecular diversity comes into play via post-synthase modification. Examples of these activities include macrocyclization, heterocyclization, aromatization, methylation, oxidation, reduction, halogenation, and glycosylation. Finally, a third level of diversity can occur in which molecules from disparate secondary metabolic pathways may interact, such as the modification of a natural product by an isoprenoid oligomer. Here, we will cover only a small subsection of... 

The modification of amino acids in proteins and peptides by oxidative processes plays a major role in the development of disease and in aging (Halliwell and Gutteridge, 1989, 1990 Kim et al., 1985 Tabor and Richardson, 1987 Stadtman, 1992). Tissue damage through free radical oxidation is known to cause various cancers, neurological degenerative conditions, pulmonary problems, inflammation, cardiovascular disease, and a host of other problems. Oxidation of protein structures can alter activity, inhibit normal protein interactions, modify amino acid side chains, cleave peptide bonds, and even cause crosslinks to form between proteins. 

Dissolve the peptide containing an N-terminal serine or threonine group at a concentration of at least 2mg/ml in 0.04 M sodium phosphate, pH 7.0. Higher concentrations of peptides or proteins may be used without modification to the rest of the protocol, because the amount of periodate used in the reaction is in sufficient molar excess, even when low-molecular-weight peptides are being oxidized. Peptides that are initially insoluble... 

Transition metals such as iron can catalyze oxidation reactions in aqueous solution, which are known to cause modification of amino acid side chains and damage to polypeptide backbones (see Chapter 1, Section 1.1 Halliwell and Gutteridge, 1984 Kim et al., 1985 Tabor and Richardson, 1987). These reactions can oxidize thiols, create aldehydes and other carbonyls on certain amino acids, and even cleave peptide bonds. The purposeful use of metal-catalyzed oxidation in the study of protein interactions has been done to map interaction surfaces or identify which regions of biomolecules are in contact during specific affinity binding events. 

Geoghegan, K.E, and Stroh, J.G. (1992) Site-directed conjugation of nonpeptide groups to peptides and proteins via periodate oxidation of a 2-amino alcohol. Applications to modification at N-terminal serine. Bioconjugate Chem. 3, 138-146. 

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