Lysine side-chains, modification

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... 

Primary amine groups on proteins consisting of N-terminal a-amines and lysine side-chain e-amines are typically present in abundant quantities for modification or conjugation reactions. Occasionally, however, a protein or peptide will not contain sufficient amounts of available amines to allow for an efficient degree of coupling to another molecule or protein. For instance, horseradish peroxidase (HRP), a popular enzyme to employ in the preparation of antibody conjugates, only possesses two free amines that... 

Both amine and lysyl oxidases possess a single Type 2 copper center and a redox-active cofactor per subunit. As shown in Figure 18, amine oxidases contain 2,4,5-trihydroxyphenylalanine quinone (TPQ) and lysyl oxidase contains lysyl tyrosine quinone (LTQ), in which a lysine side chain has been cross-linked to TPQ. These cofactors are produced from the oxidation of a specific active-site tyrosine residue via novel self-processing events that require only copper and O2 see Metal-mediated Protein Modification). 

Distinct from the lysosomal pathway are cytosolic mechanisms for degrading proteins. Chief among these mechanisms is a pathway that includes the chemical modification of a lysine side chain by the addition of ubiquitin, a 76-residue polypeptide, followed by degradation of the ubiquitin-tagged protein by a specialized proteolytic machine. Ubiquitination is a three-step process (Figure 3-13a) ... 

The useful lysine derivatives for cytochrome c are shown in Fig. 17. Chemical modifications of the lysine side chains which remove the positive charge or replace it with a negative charge destroy the interaction with cytochrome oxidase (213-215). Results with acetylation, succinyla-tion, trinitrophenylation, and guanidination are given in Table XIV. Suc-... 

Fig. 17. Chemical modifications of lysine side chains which have proved useful in studying cytochrome c mechanisms. See also M. J. Wimmer, M. Foster, K. T. Mo, D. L. Sawyers, and J. H. Harrison, Fed. Proc., Fed. Amer. Soc. Exp. Biol. Abstr. 34, 630 (1975).

Histidine, glutamate, aspartate, and lysine side chains accessible on the protein surface have been used as sites for modification by redox-active metal species. The original modification procedure involved direct reaction... 

Evidence was obtained for the presence of lysine side chains in active sites of rabbit antibodies directed to the p-azobenzoate, p-azobenzenearsonate, and p-azobenzenephosphonate groups (5). The minimum fractions of active sites containing lysine, in the three antibody populations, were 15, 45, and 25%, respectively. Maleic anhydride was used for the chemical modification. Conditions were chosen so as to minimize reactions with tyrosine, serine, and threonine. The values cited for percentages of active sites specifically modified were obtained from data on maximum hapten-binding capacity per molecule of antibody before and after treatment. Specific modification was taken as the difference between the numbers of sites inactivated in the presence of absence of a protective hapten. 

Freedman et al. (6) were also able to show that anti-p-azoben-zenearsonate antibodies can be fractionated into two separate populations having arginine or lysine side chains, respectively, in their active sites. Lysine residues were first reacted with maleic anhydride without protecting hapten present this allowed modification of lysine groups in... 

---