Carboxy biotin

The mechanism of the C02 transfer reaction with acetyl CoA to give mal-onyl CoA is thought to involve C02 as the reactive species. One proposal is that loss of C02 is favored by hydrogen-bond formation between the A -carboxy-biotin carbonyl group and a nearby acidic site in the enzyme. Simultaneous deprotonation of acetyl CoA by a basic site in the enzyme gives a thioester eno-late ion that can react with C02 as it is formed (Figure 29.6). 

Decarboxylation of /V-carboxy-biotin gives C02 plus biotin. 

Biotin reacts with an oxidized carbon fragment (denoted as CO2) and an energy-rich compound, adenosine triphosphate (ATP), to form carboxy biotin, which is activated carbon dioxide. Biotin is firmly bound to its enzyme protein by a peptide linkage. Biotin and carboxy biotin are ... 

The reactive mtermediate is l-AT-carboxy-biotin (see Figure 11.1) bound to a lysine residue of the enzyme as biocytin, which is formed from enzyme-bound biocytin by reaction with bicarbonate. 

In the bacterial biotin-dependent decarboxylases, reaction 2 proceeds from right to left, followed by decomposition of the carboxy-biotin to biotin and CO2. 

FIGURE 9,32 Structure of biotin. Biotin is used as the carrier of carboxyl groups in carboxyialion reactions. In an ATl -dcpendcnt reaction, bicarbonate is transferred to biotin, generating carboxy-biotin. The carboxyl group is bound to the I -nitrogpn. The second half of biotin-dependent carboxylation reactions involves transfer of the carboxyl group to the substrate. 

Decarboxylation of A/-carboxy-biotin gives CO2 plus biotin. 

Mechanisms of biotin-dependent reactions invariably involve two separate sequences and two separate enzyme sites, with covalently bound biotin or carboxy-biotin moving on a swinging arm from one site to the other. 

In the second step, carboxy phosphate reacts with biotin to form A-carboxy-biotin, which is the final product of the first sequence [Eq. (13)]. 

Apart from the true Claisen condensations discussed in the previous section in which the electrophilic reaction partner is another thioester, a number of enzymes also catalyze related Claisen-like condensations in which an acyl-CoA-based nucleophile reacts with other electrophilic carbonyl groups such as ketones, aldehydes, and the carboxylate group of carboxy-biotin. The most important examples of such enzymes are hydroxymethylglutaryl-CoA (HMG-CoA) synthase, citrate and homocitrate synthase (HCS), malate and ct-isopropylmalate synthase (ct-IPMS), and the biotin-dependent acetyl- and propionyl-CoA carboxylases. 

The CO 2 attached to the nitrogen of biotin is the active form of carbon dioxide which participates in numerous carboxylation reactions (e.g. acetyl-CoA to raalonyl-CoA, Chapt. XII-6, and related reactions). The very labile carboxy-biotin has now been isolated as a methyl ester (Lynen and co-workers). 

Lyases and Synthases are enzymes which catalyze the cleavage of a compound into two fragments or, in reverse, which catalyze the joining of two substances to form a third (synthases). The latter reaction is frequently equivalent to a group transfer. While it is still possible to draw up boundaries for the classification of the enzymes in this respect, for the coenzymes it ceases to be possible. Numerous groups activated by coenzymes participate in the reactions of synthases, as e.g. acetyl-CoA, carboxy-biotin, and thiamine-bound active aldehyde. 

Acetyl-CoA was found to add CO2 with the aid of a specific, biotin-containing enzyme to form malonyl-CoA. The reaction is ATP-dependent and passes through the intermediate active carboxyl, the carboxy-biotin (cf. Chapt. VI-5). Malonyl-CoA contains an unusually reactive CHj group that combines easily with acyl-CoA (acetyl-CoA or higher activated fatty acids). The result is, with loss of CO, a /3-keto acid (as shown on pagei222 ) ... 

---