Protein reaction with aldehyde

Glutaraldehyde is the most popular bis-aldehyde homobifunctional cross-linker in use today. However, a glance at glutaraldehyde s structure is not indicative of the complexity of its possible reaction mechanisms. Reactions with proteins and other amine-containing molecules would be expected to proceed through the formation of Schiff bases. Subsequent reduction with sodium cyanoborohydride or another suitable re-ductant would yield stable secondary amine linkages (Chapter 2, Section 5.3, and Chapter 3, Section 4). This reaction sequence certainly is possible, but other cross-linking reactions also are feasible. 

There are, of course, many carbonyl compounds formed by hydrolytic or oxidative deteriorations of lipid constituents, and most of these are potentially capable of entering into Mai Hard reactions with proteins. One such product is reputedly malon-aldehyde (26) (Figure 10). 

Xanthoxin is an advanced biosynthetic precursor of ABA and thus its quantification in higher plants is of interest. The aldehyde group is readily coupled directly to carrier proteins via reduction of a Schiff s base formed by reaction with protein amino-groups (Fig 3). Antiserum to such a conjugate has been produced and shows specificity for compounds having the same functionality on the cyclohexane ring [73]. Phaseic acid is a metabolite of abscisic acid and specific monoclonal antibodies have been produced to the... 

Oxidation products of Hpids (hydroperoxides, free alkoxyl and peroxyl radicals, epoxides and aldehydes) react with a number of food constituents during the processing and storage of food. These reactions often lead to a reduction in the nutritional value of foods (such as reactions with proteins and vitamins) and a deterioration of their organoleptic properties (e.g. reactions with flavour active substances). 

The N-terminal methionine residue of protein can also be employed for selective PEGylation using aldehyde-terminated PEG via a reductive amination reaction, because the N-terminal primary amine has a lower pAa of 7.8 than other amines such as lysines, whose pZa is 10.1 [7]. After reaction with aldehyde-terminated PEG at low pH, the resultant imine is reduced with sodium cyanoborohydrate to provide PEGylated protein (Fig. 4) [8, 9]. This technique was used for the production of Neulasta, which was approved for use by the FDA in 2002 [10]. 

Fig. 4 PEGylation at the N-terminal methionine residue. The difference in pKa. between the N-terminal amine and other amines in the protein enables site-specific PEGylation. After reaction with aldehyde-terminated PEG at low pH, reduction of the resultant imine produces PEGylated protein...

Aldehyde particles are spontaneously reactive with hydrazine or hydrazide derivatives, forming hydrazone linkages upon Schiff base formation. Reactions with amine-containing molecules, such as proteins, can be done through a reductive amination process using sodium cyanoborohydride (Figure 14.21). 

In a fume hood, add 10 pi of 5M sodium cyanoborohydride (Sigma) per ml of reaction solution. Caution Cyanoborohydride is extremely toxic. All operations should be done with care in a fume hood. Also, avoid any contact with the reagent, as the 5M solution is prepared in IN NaOH. The addition of a reductant is necessary for stabilization of the Schiff bases formed between an amine-containing protein and the aldehydes on the antibody. For coupling to a hydrazide-activated protein, however, most protocols do not include a reduction step. Even so, hydrazone linkages may be further stabilized by cyanoborohydride reduction. The addition of a reductant during hydrazide/aldehyde reactions also increases the efficiency and yield of the reaction. 

By Far the simplest bifunctional crosslinking agent is formaldehyde. Although structurally a mono-functional aldehyde compound, formaldehyde reacts with proteins via a two-step reaction... 

For this reaction, soluble monomers are needed, e.g. a mixture of N AT-methylene bisacrylamide as crosslinker, methacrylamide as an inert comonomer, methacrylic acid as ionic comonomer for stabilization [309] and methacryl ami-do-AT-acetaldehyde-dimethylacetal as functional comonomer. The coupling with proteins is only possible if the free aldehyde groups are accessible, i.e. if they are not located in the interior of the microgel. This condition can only be fulfilled by a careful choice of the comonomer composition in the reaction mixture [291]. 

Photochemical cis-trans isomerization in a conjugated polyene system is thought to be the crucial primary process in vision. The visual pigment (rhodopsin) is derived from 11 -crs-retinal by reaction of the aldehyde group with an amino substituent in a protein (opsin). There is considerable distortion in the geometry of this chromophoric group anyway, because of the spatial requirements of the protein... 

---