Affinity labeling, chemical modification

As with chemical affinity labeling, the modification of an amino acid may interfere with protein fragmentation reactions, i.e., the Edman degradation, and cyanogen bromide cleavage. ... 

Fig. 4. Classification of reported noncompetitive immunoassays for haptens based on the assay principle. (A) Assays that include a chemical modification of hapten to allow sandwich-type detection. (B1) Improved single-antibody immunometric assays that separate immune complex and excess labeled antibody, either by using a hapten-immobilized affinity column or based on differences in their physical properties. (B2) A variation of single-antibody immunometric assays based on masking of unoccupied antibody by an immunoreactive macromolecule followed by selective capture and detection of the hapten-occupied antibody. (C) Assays employing a probe molecule specific to a hapten-antibody complex.

An affinity label, or active-site-directed irreversible inhibitor, is a chemically reactive compound that is designed to resemble a substrate of an enzyme, so that it binds specifically to the active site and forms covalent bonds with the protein residues.1-3 Affinity labels are very useful for identifying catalytically important residues and determining their pKa values from the pH dependence of the rate of modification. 

Affinity labeling and chemical modification studies were carried out with purified Fi. The /3Thr-287, /3Ile-290 and /3Tyr-297 residues ( coli numbering) of beef heart Ft were... 

Affinity Labeling. As described in the previous section, differences in the reactivity to chemical modification of groups of a single type can be interpreted in terms of differences in their locations and environments within the protein molecule. A similar approach, but one that probes... 

Chemical modification can be used to obtain information on the catalytic mechanism and on the catalytic site of the enzyme of interest. One goal in the design of affinity labels for enzymes is to determine the catalytically important residues. First, the affinity label has to behave as an analogue of the substrate (or of the activator or inhibitor) by competition experiments. 

Chemical modification studies on ADPGlc PPase have involved the use of the following affinity labels ... 

---