Succinic reaction with protein

Reactions with succinic anhydride or acetic anhydride to block dendrimer amines can be done in aqueous or methanolic solution. If organic solvent is used for the reaction, then it is typical to include triethylamine as a proton acceptor, which helps drive the reaction. Such reactions, however, can t be done to dendrimer amines once a protein containing amines also has been conjugated, as the protein too will get modified. 

Habeeb, A.F.S.A., Cassidy, H.G., and Singer, S.J. (1958) Molecular structural effects produced in proteins by reaction with succinic anhydride. Biochim. Biophys. Acta 29, 587. 

Acid anhydrides, as their name implies, are formed from the dehydration reaction of two carboxylic acid groups (Fig. 72). Anhydrides are highly reactive toward nucleophiles and are able to acylate a number of the important functional groups of proteins and other macromolecules. Upon nucleophilic attack, the anhydride yields one carboxylic acid for every acylated product. If the anhydride was formed from monocar-boxylic acids, such as acetic anhydride, then the acylation occurs with release of one carboxylate group. However for dicarboxylic acid anhydrides, such as succinic anhydride, upon reaction with a nucleophile the ring structure of the anhydride opens, forming the acylated product modified to contain a newly formed carboxylate group. 

The action of trypsin on proteins and peptides can be limited to the bonds formed by arginine if the e-ammonium groups of lysine are modified. Reaction of proteins with fluorodinitrobenzene (Anfinsen and Redfield, 1956), carbobenzoxychloride (Anfinsen et al., 1956), 0-methylisourea (Weil and Talka, 1957), succinic anhydride (Li and Bertsch, 1960), potassium cyanate (Stark and Smyth, 1963), carbon disulfide (Merigan et al.,... 

This class of haptens includes those that have a carboxyl group, such as acetylsalicylic acid (aspirin) or the peptides angiotensin and brady-kinin. In addition, many haptens, such as some steroids, may have reactive groups to which a carboxyl group can be attached as, for example, by reaction with succinic anhydride (see below). For conjugation to proteins, the same procedures may be used regardless of whether the carboxyl group is present as an inherent part of the hapten or as an added moiety. 

NEM is not completely specific for sulfhydryl groups. Reaction with amino groups occurs readily in native proteins. Formation of such derivatives would be detected by a comparison of the yield of S-succinylcysteine and ethylamine, or by the appearance of N-e-succin-2-yl-lysine, or N-succin-2-yl-histidine (Holbrook and Jeckel 1969) upon acid hydrolysis of the modified protein (cf. ch. 2). 

Maleic anhydride, 2-methylmaleic anhydride (citraconic anhydride), and 2,3-dimethyImaleic anhydride react with proteins in a manner similar to succinic anhydride, but the products are much more labile to hydrolysis [5,6]. The reaction products of maleic anhydride and amino groups are stable at neutral pH but rapidly hydrolyze when acidified to pH 3.5. This lability increases in the order maleyl < citraconyl < dimethylmaleyl derivatives. Citraconylamides are stable enough to withstand many hours at neutral or alkaline pH s but... 

So, the biosynthesis of methionine (Met, M), the first of the essential amino adds to be considered (Scheme 12.13), begins by the conversion of aspartate (Asp, D) to aspartate semialdehyde in the same way glutamate (Glu, E) was converted to glutamate semialdehyde (vide supra. Scheme 12.6). Phosphorylation on the terminal carboxylate of aspartate (Asp, D) by ATP in the presence of aspartate kinase (EC 2.7.2.4) and subsequent reduction of the aspart-4 yl phosphate by NADPH in the presence of aspartate semialdehyde dehydrogenase (EC 1.2.1.11) yields the aspartate semialdehyde. The aspartate semialdehyde is further reduced to homoserine (homoserine oxoreductase, EC 1.1.1.3) and the latter is succinylated by succinyl-CoA with the liberation of coenzyme A (CoA-SH) in the presence of homoserine O-succinyl-transferase (EC 2.3.1.46). Then, reaction with cysteine (Cys, C) in the presence of cystathionine y-synthase (EC 2.5.1.48) produces cystathionine and succinate. In the presence of the pyridoxal phosphate protein cystathionine P-lyase (EC 4.4.1.8), both ammonia and pyruvate are lost from cystathionine and homocysteine is produced. Finally, methylation on sulfur to generate methionine (Met, M) occurs by the donation of the methyl from 5-methyltetrahydrofolate in the presence of methonine synthase (EC 2.1.1.13). 

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