CHEMICAL PROPERTIES 



341 



transamidation to form a thiazane derivative. The product of the reac- 

 tion of cysteine and the colored iV-2,4-dinitroanilinomaleimide undergoes 

 cyclization, involving the amino group of cysteine and resulting in scission 

 of the succinimide ring, the final product being a substituted 1,4-thiazine. 

 O 



iV- (4 -Hydroxy- 1- 

 naphthyl) maleimide 



7V-(4-Diniethylamino- 

 3, 5-dinitrophenyl) 

 maleimide 



W // 



NO, 



O NO, 



A^-2,4-Dinitro- 

 anilinomaleimide 



It has been generally supposed that A'^-ethylmaleimide is specific for SH 

 groups, since no reaction with cystine, oxidized glutathione, and various 

 amino acids had been detected, but Smyth et al. (1960) have shown that 

 under certain conditions (200 milf of reagents and pH 6.4-7.4) imidazole 

 and some amino acids are iV-acylated by iV-ethylmaleimide. However, in 

 the case of imidazole, there is subsequent hydrolysis to iV-ethylmaleamate, 

 which may be further acylated to initiate polymerization. Histidine and 

 some histidine peptides also give this reaction. It is not known if such 

 acylations are of importance in the reactions of iV-ethylmaleimide with pro- 

 teins and enzymes, but it is always possible that certain histidine residues 

 or amino groups on proteins may be in a particularly reactive state. When 

 glycylalanine is incubated with 10 mM iV-ethylmaleimide at pH 7.4 and 

 37° for 2 hr, the amino group adds to the double bond without opening 

 of the imide ring (Smyth et al, 1964). The amino groups of most proteins 

 do not react so readily as shown by comparing the yields of ethylamine 

 (formed from reaction with amino and SH groups) and *S-(l,2-dicarboxy- 

 ethyl)-L-cysteine (formed from reaction with SH groups) from the hydrol- 

 ysis of proteins previously treated with A''-ethylmaleimide. It is pointed 

 out that specificity toward protein SH groups is favored by a pH below 

 neutrality and avoiding excess reagent, but that each protein or enzyme 

 must be treated as a special problem. 



