Carbohydrates reducing ends

Another form of post-translational modification that may add carbohydrate to a polypeptide is non-enzymatic glycation. This reaction occurs between the reducing ends of sugar molecules and the amino groups of proteins and peptides. See Section 2.1 in this chapter for further details and the reaction sequence behind this modification. 

Amine functionalities also may be created on polysaccharides (Section 4.3, this chapter). The reducing ends of carbohydrate molecules (or generated aldehydes) may be reacted with small diamine compounds to yield short alkylamine spacers that can be used for subsequent conjugation reactions. Hydrazide groups may be similarly created using f z s-hydrazide compounds (Sections 4.5 and 4.6, this chapter). 

Retention of native carbohydrate structure also is important in applications that utilize the conjugated polysaccharide in binding studies with receptors or lectins. In these cases, the carbohydrate should be modified at limited sites, preferentially only at its reducing end. Section 4.6 of this chapter discusses glycan conjugation techniques in greater detail. 

In some instances, reducing sugars are present that can be reductively aminated without prior periodate treatment. A reducing end of a monosaccharide, a disaccharide, or a polysaccharide chain may be coupled to a diamine by reductive amination to yield an aminoalkyl derivative bound by a secondary amine linkage (Figure 1.96). Also see Section 4.6, this chapter, for an extensive discussion on carbohydrate modification techniques. 

Figure 1.117 Glucosylamine derivatives can be prepared at the reducing end of glycans or other reducing carbohydrates by reaction with ammonium carbonate. The resultant amine derivative can be used to conjugate the carbohydrate with other proteins or molecules without disturbing the cyclic character of the reducing end.

Figure 7.15 The reducing end of a glycan or a carbohydrate can be used to conjugate to an amine-dendrimer by reductive amination, which results in the formation of a secondary amine linkage.

Dissolve an oligosaccharide or glycan having a reducing end to be modified in 2 1 pyri-dine/glacial acetic acid (vol/vol) with a total reaction volume of 10-100 pi. If the carbohydrate initially is insoluble in the reaction solution, a prior dissolution in a minimal amount of DMSO or water can be done and then an aliquot transferred to the reaction medium. 

In some cases, the ability to modify glycans at the reducing end without reduction preserves the carbohydrate s native structure sufficiently to allow interactions with proteins that would otherwise not interact if the bond were reduced. Therefore, depending on the ultimate use of the biotinylated carbohydrate, using a hydrazide mediated conjugation process can have advantages over the use of amine-biotin compounds. 

Figure 18.19 Biotin-PEG4-hydrazide is a hydrophilic biotinylation reagent that can be used to modify glycans or carbohydrates at their reducing end or after periodate oxidation to create aldehydes.

In one scenario, the first carbohydrate is anchored via its reducing end (see Scheme 2.1, case 1). Here the support-bound carbohydrate will function as an acceptor in the coupling step to a solution-based donor D. As the next cycle is contemplated, a unique acceptor hydroxyl must be exposed in the now elongated, resin-bound carbohydrate constmct. 

FIGURE 5.1 Cluster model of amylopectin. A and B denote nomenclature of branch chains, 0=reducing end, c.l. = chain length in degree of polymerization. Reprinted from Carbohydrate Research, Vol. 147, Hizukuri (1986), Polymodal distribution of the chain lengths of amylopectin, and its significance, Pages 342-347, with permission from Elsevier. 

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