Carbamylation reactions

C rb myl tion. Modification of the amino-terminal groups of hemoglobin (Hb) by the carbamylation reaction using isocyanic acid [75-13-8]... 

Rubisco exists in three forms an inactive form designated E a carbamylated, but inactive, form designated EC and an active form, ECM, which is carbamylated and has Mg at its active sites as well. Carbamylation of rubisco takes place by addition of COg to its Lys ° e-NHg groups (to give e—NH—COO derivatives). The COg molecules used to carbamylate Lys residues do not become substrates. The carbamylation reaction is promoted by slightly alkaline pH (pH 8). Carbamylation of rubisco completes the formation of a binding site for the Mg that participates in the catalytic reaction. Once Mg binds to EC, rubisco achieves its active CM form. Activated rubisco displays a Ai, for CO2 of 10 to 20... 

In vivo intermediate in mammals by the detection of two of its derivatives,, the glutathione (GSH) conjugate and its further metabolites formed by an initial carbamylation reaction (W) (see below) and 2-chloroacrolein detected in the microsome-NADPH system and derived from the rearrangement-elimination reaction sequence discussed above (6). Sulfallate also yields 2-chloroacrolein in the microsome-NADPH system, presumably by -CH2 hydroxylation (22) on analogy with the metabolism of EPTC shown previously. 

Carbamylation Reactions. -Alkyl, -benzyl and -chloroallyl thiocarbamates do not readily react with GSH. In contrast, their sulfoxide derivatives ( and 6,) are very effective carbamylating agents for many thiols including GSH (19, ). The GSH conjugates formed vivo via 3 and 6 are quickly cleaved, acetylated and further metabolized as follows (19-21. 23. 24). 

Before substrate binding can take place, rubisco must first be activated. This occurs via carbamylation (reaction with CO2) of an essential Lys residue . This promotes the binding of an essential Mg + ion after which the active site is complete. Rubisco can now recognize and bind the first substrate which is ribulose-P2 (D-ribulose 1,5-bisphosphate) . The substrate is bound to the Mg + ion via an inner-sphere coordination of the C2-carbony 1 and C3-hydroxyl groups which appropriately positions and activates the ribulose-P2 for subsequent reaction. Substrate binding causes a flexible loop to close over the active site which buries the active site deep within the protein and restricts access to a small channel just large enough for CO2 (and 02) . 

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... 

Intermediates in elimination reactions could also be detected using this three-phase test on the carbamylation reaction (R = NHCeH in resin 9). The reaction was catalyzed by triethylamine or a proton sponge [1,8-bis(dimethylamino)naphthalene]. The intermediate, phenyl isocyanate, was readily observed (infrared spectra) between the two resins and was trapped as the urea derivative of the resin (11). 

Ketone formation can also be avoided if one of the functional acyl halogens ia phosgene is blocked. Carbamyl chlorides, readily obtained by the reaction of phosgene with ammonia or amines, are suitable reagents for the preparation of amides ia direct Friedel-Crafts acylation of aromatics. The resulting amides can be hydroly2ed to the corresponding acids (134) ... 

The practical appHcation of this reaction has been demonstrated ia the preparation of terephthaUc acid from toluene, ia which case oxidation follows hydrolysis (135). The reaction also proceeds well with substituted carbamyl chlorides such as CgH (C2H )NCOCl. 

In these reactions the active acylating agent is the carbamyl chloride, formed by the reaction of the isocyanate with hydrogen chloride (137) ... 

The most important commercial process is the reaction of MDA with an excess of phosgene to form the corresponding isocyanate, 4,4 -methylene-diphenyldiisocyanate, MDI, C H qN202- The reaction proceeds through the formation of a primary carbamyl chloride that is decomposed with heating and the removal of HQ. 

Substrate RuBP binds much more tightly to the inactive E form of rubisco (An = 20 nM) than to the active ECM form (A, for RuBP = 20 ixM). Thus, RuBP is also a potent inhibitor of rubisco activity. Release of RuBP from the active site of rubisco is mediated by rubisco activase. Rubisco activase is a regulatory protein it binds to A-form rubisco and, in an ATP-dependent reaction, promotes the release of RuBP. Rubisco then becomes activated by carbamylation and Mg binding. Rubisco activase itself is activated in an indirect manner by light. Thus, light is the ultimate activator of rubisco. 

In this scheme, EOH is the enzyme, IX is the inhibitor (either a carbamate or an organophosphate). EOH(IX) is analogous to the Michaelis Menton comploc seen with the substrate reaction. EOI is the acyl-enzyme intermediate for carbamates or a phosphoro-enzyme intermediate for the organophosphates. The equilibrium constant for this reaction (K ) is defined as k /k and the phosphorylation or carbamylation constant is defined as k2- In this study 42)y ANTX-A(S) was found to be more specific for AChE than BUChE. The double reciprocal and Dixon plot of the inhibition of electric eel AChE indicated that the toxin is a non-competitive inhibitor decreases, k remains unchanged) (Figure 2). 

Reactions involving carbamyl phosphate in the degradation of arginine in Clostridia, and the fermentation of allantoin by Streptococcus allantoicus... 

Carbamyl phosphate condenses with ornithine to yield citrulline in the ornithine transcarbamylase (OTC) reaction. OTC is encoded on band p21.1 of the X chromosome, where the gene contains 8 exons and spans 85 kb of DNA. The activity of this enzyme is directly related to dietary protein. There may be tunneling of ornithine transported from the cytosol to OTC, with the availability of intramitochondrial ornithine serving to regulate the reaction. 

A phosphotriesterase isolated from the soil bacterium Pseudomonas diminuta is the best characterized enzyme of this type. There is evidence for the presence of two active site Zn2+ ions in vivo. A crystal structure of the dinuclear Cd2+ form is available in which the metal ions are bridged by a carbamylated Lys-amino group with a metal-metal distance of 3.8 A [ 18]. Substrate hydrolysis follows a SN2 type reaction and nucleophilic attack of M-OH is likely, but mechanistic details are not yet clear. 

PITC has been used extensively in the sequencing of peptides and proteins and reactions under alkaline conditions with both primary and secondary amino acids. The methods of sample preparation and derivatization follow a stringent procedure which involves many labour-intensive stages. However, the resulting phenylthio-carbamyl-amino acids (PTC-AA s) are very stable, and the timing of the derivatization step is not as critical as when using OPA. 

Reaction of alkyl or aryl isocyanates with benzimidazolinethione (254) affords the iV-carbamyl derivative (255). Treatment of (255) with bromine in triethylamine gives the benzimidazolo[l,2-r/]-1,2,4-thiadiazoline (256) (Scheme 58). A related thioanalogue (258) was obtained by a similar process from the A-thiocarbomyl derivative (257) (Equation (35)) <83X2311). 

Reaction of Carbamyl Chlorides and Derivatives with Hydrazines... 

Carbamates, transesteriflcation, 243 IV-Carbamates, 223-234 O-Carbamates, 235-245 Carbamic acid derivatives, reactions to give semicar bazides, 198-199 Carbamoyl chloride, 244 Carbamyl chlorides, 198-200 Carbanilide, see jym-Diphenylurea Carbenes, 52, 62... 

Ureas, 134-165 disubstituted, 148 monosubstituted, 148 0-alkylation, 178-179 prepared via oxidative carbamylation, 158 reaction with alcohols, 236-239 with amines, 135-138 tetrasubstituted, 145-146 Urethanes, see iV-Carbamates... 

Carbamyl chlorides, available by the reaction of phosgene on secondary amines, can react with hydrazines to give good yields of 1,1-dialkylsemi-carbazides [31] (Eq. 24). 

Examples of the products of the reaction of carbamyl chlorides and derivatives with hydrazines are listed in Table VIII. 

Pandey, V.N. Pradhan, D.S. Reverse and forward reactions of carbamyl phosphokinase from Streptococcus faecalis R. Participation of nucleotides and reaction mechanisms. Biochim. Biophys. Acta, 660, 284-292 (1981)... 

Reactions between 5-cyanotetrazole and transition metals, when performed in boiling acetone, lead to hydrolysis of the cyano group and formation of 5-carbamyl tetrazolate complexes (68). Complexes containing 1- or 5-substituted tetrazolate anions can also be obtained by 1,3-dipolar cycloaddition of organic isonitriles (RNC) (15) or nitriles (RCN) (61), respectively, to coordinated azide ligands [Eqs. (3) and (4)]. 

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