Tandem chemical modification

Figure 2.7. Identification ofphosphoproteins by site-specific chemical modification. A. Method of Zhou et al. (2001) involves trypsin digest of complex protein mixture followed by addition of sulfhydryl groups specifically to phosphopeptides. The sulfhydryl group allows capture of the peptide on a bead. Elution of the peptides restores the phosphate and the resulting phosphopeptide is analyzed by tandem mass spectrometry. B. Method of creates a biotin tag in place of the phosphate group. The biotin tag is used for subsequent affinity purification. The purified proteins are proteolyzed and identified by mass spectrometry.

Wallace A., Boyce J., and Balland A. (2002), Structural Characterization of Man-nose-6-phosphate-containing glycans by tandem mass spectrometry, IBC s Conference on The Impact of Post-Translational and Chemical Modifications on Protein Therapeutics, San Diego, CA. 

Liu and Hop [19] have reviewed various LC-tandem MS strategies, with or without chemical modification or derivatization, which have been successfully applied in the identification and the rational, but tentative, structural determination of drug metabolites. These techniques are equally applicable to the analysis of known plant secondary metabolites or to the de novo structural identification of new compounds. 

Microderivatization or chemical modification of analytical samples can be useful in enhancing the ion response, and hence the LC-MS sensitivity, for any particular analyte. In the case of alkaloids, which are already capable of ready ionization, chemical modification can be more valuable in elucidating structures by providing tandem MS evidence for positions of substitution in the parent molecule. Such derivatization or chemical modification for HPLC-MS can include H/D exchange by using deuterated mobile phases, Jones oxidation of aliphatic hydroxyls, selective acetylation of hydroxyl and amine groups, and N-oxide reduction [19,20]. 

The Squibb team also worked back from their conception of the receptor s shape, based on their knowledge of details of the cleavage reaction it performed. Since the ACE protein had not itself been structurally defined, they developed a hypothetical model based on the active site of another enzyme that performed a similar cleavage reaction, and whose structure was known from x-ray crystallography. The research team crafted compounds to fit into the hypothetical active site of ACE and discovered a compound with inhibitory activity that was similar in shape to two amino-acids in tandem. They performed further chemical modifications of this molecule and eventually found a chemical amenable to oral administration that was 1,000 times more potent than their initial lead molecule. The synthetic drug had greater activity than the nine-amino-acid venom peptide. 

Menlyadiev, M. Stone, J.A. Eiceman, G.A., Tandem ion mobility measurements with chemical modification of ions selected by compensation voltage in differential mobility spectrometry/differential mobifity spectrometry instrument, Int. J. Ion Mobil. Spectrom. 2012,15,123-130. 

Conrotto, R Heilman, U. Lys Tag An easy and robust chemical modification for improved de novo sequencing with a matrix-assisted laser desorption/ionization tandem time-of-flight mass spectrometer. Rapid Commun. Mass. Spectrom. 2008, 22,... 

We had earlier deduced the total amino acid sequences of some unknown blue-green algal toxic peptides utilizing tandem mass spectrometric techniques in combination with simple chemical modifications and derivatization procedures. The cyclic peptides were hydrolyzed using trifluoroacetic acid. The linear hydrolytic products and their corresponding methyl esters were subjected to FAB ionization and the individual types of ions were analyzed using CID processes. The sequences of the individual peptides were derived from the corresponding recorded MS/MS spectra. Some of the peptides were found to contain Asp and dehydroalanine which had not been observed previously. 

Cesium Separation. Cesium will be removed from the waste super-nate by sorption on a phenol-sulfonic ion exchange resin such as Duolite (Diamond Shamrock Chemical Co.) ARC 359, as shown in Figure 3. This flowsheet is a modification of one currently being used by ARHCO at Hanford (3), Cesium will be absorbed on the two columns in tandem until breakthrough from the first column exceeds a predetermined level, after which the column will be washed with water (not shown in the diagram) and eluted with a mixture of ammonium carbonate and ammonium hydroxide. Breakthrough will be detected by a gamma ray monitor on the line between the two columns. 

It is worth noting that the radical damage to methionine-containing peptides and proteins consists of a desulfurization process, which leads to the replacement of a methionine residue with an a-aminobutyric acid in the sequence. This could be a posttranslational modification, which is linked to a postsynthetic modification of lipids by the above-reported tandem mechanism. A chemical biology approach can be proposed involving lipidomics and proteomics, in order to configure the metabolic changes related to a radical stress. 

The main oxidation reactions of the 2-deoxyribose of DNA are mediated by "OH that are able to abstract hydrogen atoms from most of the positions with the exception of the methylene group at C2, which is a poorly reactive site [12, 14, 112], An abundant literature is available on the degradation pathways that are derived from the reactions of osidic carbon-centered radicals and that lead in most cases to the formation of strand breaks [12, 14, 112], However, there is one major exception that concerns the chemical reactions of the CT radical that is the precursor of 2-deoxyribonolactone [113]. Here, emphasis is placed on reactions of C4 and C5 radicals that may lead to the formation of base modifications either as tandem lesions or clustered damage. 

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