Lysine reaction with carbon dioxide

Concerns about effects of supercritical carbon dioxide on food quality has prompted related investigations on the possible reactions of amino acids with carbon dioxide and water. Weder found that moist supercritical carbon dioxide at 80 °C did not react with any of the examined amino acids including lysine. Weder also found that Lysozyme lipase unfolded and partially oligomerized in moist SCCO2. He concluded that denaturation was caused by heating the protein in the presence of water and not by interactions with SCCO2 [26],... 

Biotin functions to transfer carbon dioxide in a small number of carboxylation reactions. A holocarboxylase synthetase acts on a lysine residue of the apoenzymes of acetyl-CoA carboxylase, pymvate carboxylase, propi-onyl-CoA carboxylase, or methylcrotonyl-CoA carboxylase to react with free biotin to form the biocytin residue of the holoenzyme. The reactive intermediate is 1-7V-carboxybiocytin, formed from bicarbonate in an ATP-dependent reaction. The carboxyl group is then transferred to the substrate for carboxylation (Figure 21—1). 

The way biotin participates in carbon dioxide fixation was established in the early 1960s. In 1961 Kaziro and Ochoa using propionyl CoA carboxylase provided evidence for 14C02 binding in an enzyme-biotin complex. With excess propionyl CoA the 14C label moved into a stable position in methyl malonyl CoA. In the same year Lynen found biotin itself could act as a C02 acceptor in a fixation reaction catalyzed by B-methylcrotonyl CoA carboxylase. The labile C02 adduct was stabilized by esterification with diazomethane and the dimethyl ester shown to be identical with the chemically synthesized molecule. X-ray analysis of the bis-p-bromanilide confirmed the carbon dioxide had been incorporated into the N opposite to the point of attachment of the side chain. Proteolytic digestion and the isolation of biocytin established the biotin was bound to the e-NH2 of lysine. 

Biotin (5) is the coenzyme of the carboxylases. Like pyridoxal phosphate, it has an amide-type bond via the carboxyl group with a lysine residue of the carboxylase. This bond is catalyzed by a specific enzyme. Using ATP, biotin reacts with hydrogen carbonate (HCOa ) to form N-carboxybiotin. From this activated form, carbon dioxide (CO2) is then transferred to other molecules, into which a carboxyl group is introduced in this way. Examples of biotindependent reactions of this type include the formation of oxaloacetic acid from pyruvate (see p. 154) and the synthesis of malonyl-CoA from acetyl-CoA (see p. 162). 

This FAD-dependent enzyme [EC 1.13.12.2] catalyzes the reaction of L-lysine with dioxygen to produce 5-aminopentanamide, carbon dioxide, and water. Other diamino acids can serve as substrates as well. 

This enzyme catalyzes the decarboxylation of acetoacetate to acetone and carbon dioxide. The nonenzymatic reaction involves the expulsion of a highly basic eno-late ion at neutral pH (equation 2.36), but the enzymatic reaction circumvents this by the prior formation of a Schiff base with a lysine residue. The protonated imine is then readily expelled. 

The important function of biotin is its role as coenzyme for carboxylase, which catalyses carbon dioxide fixation or carboxylation reaction. The epsilon amino group of lysine in carboxylase enzymes combines with the carboxyl group of biotin to form covalently linked biotinyl carboxyl carrier protein (BCCP or biocytin) (Figure 6.8). This serves as an intermediate carrier of carbon dioxide. The carboxylation of acetyl CoA to malonyl CoA in presence of acetyl CoA carboxylase requires biotin as coenzyme. Propionyl carboxylase and pyruvate carboxylase are also associated with biotin. 

Although very interesting biotranformations have been reported in supercritical carbon dioxide, this solvent has been found to affect enzyme activity adversely. CO can react reversibly with free amino groups (lysine residues, specifically) on the surface of the protein to form carbamates, leading to low activity enzyme. [21]. Furthermore, carbon dioxide dissolves in water at molar concentrations at moderate pressures (<100 bar) and rapidly forms H COj. This can create some problems in biocatalytic reactions because many enzymes are denatured (unfolded and/or deactivated) at low pH. Enzymes can also be denatured by pressurization/depressuriza-tion cycles. For all of them, it is necessary to develop new enzyme stabilization strategies. 

The enzyme phosphotriesterase hydrolyzes many different organophosphorus triesters, including several acetylcholinesterase inhibitors. The chemical mechanism involves an activated water molecule that directly attacks the phosphorus center with a resultant inversion of configuration [708]. Thus the overall reaction mechanism does not involve the formation of a phosphorylated-enzyme intermediate, but it does involve a Schiff base-type interaction between carbon dioxide and a lysine residue giving a carbamate group in the active site. 

Biotin is a carrier of carbon dioxide it has a specific site for covalent attachment of GOg (Figure 18.7). The carboxyl group of the biotin forms an amide bond with the e-amino group of a specific lysine side chain of pyruvate carboxylase. The GOg is attached to the biotin, which, in turn, is covalently bonded to the enzyme, and then the GOg is shifted to pyruvate to form oxaloacetate (Figure 18.8). Note that ATP is required for this reaction. 

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