Carbamylation with cyanate

The chloroketone reactive sulfhydryl group in the glutamine binding site of carbamyl phosphate synthetase reacts also with cyanate. " However, the inactive cyanate-enzyme complex regains activity upon incubation at alkaline pH. An S-[ C]carbamylcysteine derivative was isolated from the [" C] cyanate-enzyme complex after digestion with Pronase. S-Carbamyl cysteine moves with 22/ = 0.17 in butanol-acetic acid-water (12 3 5, v/v). 

The synthesis of pyrophosphate under primitive earth conditions has been shown by S. L. Miller and M. Parris. > It occurs by reaction of orthophosphate with cyanate on the surface of hydroxylapatite, presumably forming carbamyl phosphate which reacts with another phosphate molecule to generate pyrophosphate. The formation of the pyrophosphate bond has also been accomplished by extension of the above reaction methods. Thus, NH4H2PO4 in the presence of urea at low humidity and below 100°C forms inorganic polyphosphate or pyrophosphate. Once inorganic pyrophosphate is available, trans-phosphorylations can take place. In this way AMP has been further phosphorylated to ADP and ATP. 

Carbamyl and thiocarbamyl fluorides are obtained from hydrogen fluoride and cyanic acid or alkali metal cyanates or thiocyanates [/, 54] Nitnles give iraidofluoride salts with hydrogen fluoride [7] whereas hydrogen cyanide affords difluoromethylamine, which can be isolated as its hexafluoroarsinate salt [55) (equation II)... 

Acetate kinase is phosphorylated by acetyl phosphate and it has been shown that the phosphoenzyme can synthesise ATP from ADP, and acetyl phosphate from acetate. The mode of decomposition of carbamyl phosphate in aqueous solution is pH dependent and can proceed with either the production of ammonia and carbon dioxide (equation 1), or cyanate (equation 2). No cyanate could be detected during the hydrolysis... 

Preparation of Chemically Modified Collagen Membranes. Collagen membrane was prepared by the above procedure. The lysyl E-amino residues of collagen, in membrane form, were modified to varying degree by reaction with potassium cyanate. The collagen membrane was carbamylated by immersing preswollen films in 0.4M potassium cyanate solution, pH 8.5 for different periods of time. The carbamylation reaction was carried out at ambient temperature (approx. 20°C). The pH of the reaction mixture was monitored with a pH meter and the pH maintained at 8.5 by the addition of 0.05N... 

The extent of modification can be assessed either by using "relabeled cyanate, or for lysine by determining the homocitrulline content of the protein after acid hydrolysis. Since homocitrulline is slowly hydrolyzed to lysine, the carbamylated protein should be hydrolyzed for 24 and 48 hr, and the values for homocitrulline extrapolated with first order kinetics to zero time (Plapp et al. 1971 2.7.4). 

Cyanate present in aqueous urea solutions reacts with the free amino and sulfhydryl groups of proteins. Urea is often tacitly assumed to be a reagent, which alters the structure of the protein and may be used to keep target proteins on their monomeric form during purification. However, at pH 6 and above urea hydrolyses under formation of cyanate leading to carbamylation reactive groups in proteins. 

Stark, G. R. Reactions of cyanate with functional groups of proteins. IV. Inertness of aliphatic hydroxyl groups. Formation of carbamyl- and acylhydantoins. Biochemistry 4 2363-2367,1965. 

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