Active intermediates enzymatic reactions

Lipases have also been used as initiators for the polymerization of lactones such as /3-bu tyro lac tone, <5-valerolactone, e-caprolactone, and macrolides.341,352-357 In this case, the key step is the reaction of lactone with die serine residue at the catalytically active site to form an acyl-enzyme hydroxy-terminated activated intermediate. This intermediate then reacts with the terminal hydroxyl group of a n-mer chain to produce an (n + i)-mer.325,355,358,359 Enzymatic lactone polymerization follows a conventional Michaelis-Menten enzymatic kinetics353 and presents a controlled character, without termination and chain transfer,355 although more or less controlled factors, such as water content of the enzyme, may affect polymerization rate and the nature of endgroups.360... 

The enzymatic reactions of peroxidases and oxygenases involve a two-electron oxidation of iron(III) and the formation of highly reactive [Fe O] " species with a formal oxidation state of +V. Direct (spectroscopic) evidence of the formation of a genuine iron(V) compound is elusive because of the short life times of the reactive intermediates [173, 174]. These species have been safely inferred from enzymatic considerations as the active oxidants for several oxidation reactions catalyzed by nonheme iron centers with innocent, that is, redox-inactive, ligands [175]. This conclusion is different from those known for heme peroxidases and oxygenases... 

A series of subsequent reactions after PAL first introduces a hydroxyl at the 4-position of the ring of cinnamic acid to form p- or 4-coumaric acid (i.e., 4-hydroxycinnamic acid). Addition of a second hydroxyl at the 3-position yields caffeic acid, whereas O-methylation of this hydroxyl group produces ferulic acid (see Fig. 3.3). Two additional enzymatic reactions are necessary to produce sinapic acid. These hy-drocinnamic acids are not found in significant amounts in plant tissue because they are rapidly converted to coenzyme A esters, or glucose esters. These activated intermediates form an important branch point because they can participate in a wide range of subsequent reactions. 

Flavin oxidation of carbanions has also been of much concern since active intermediates in some flavoenzyme-mediated reactions (amino acid oxidase, lactate oxidase, etc.) are carbanions (Kosman, 1977). Flavin oxidation of nitroethane carbanion (20), which had not been achieved in non-enzymatic systems, occurs with [56] bound to CTAB micelles (Shinkai etal., 1976b). This suggests that the nitroethane carbanion is also activated by the micellar environment. 

An enzymatic reaction intermediate formed by carboxyl group transfer to a hydroxyl group of a serine residue within the active site. 

Propionyl-CoA is first carboxylated to form the d stereoisomer of methylmalonyl-CoA (Pig. 17—11) by propionyl-CoA carboxylase, which contains the cofactor biotin. In this enzymatic reaction, as in the pyruvate carboxylase reaction (see Pig. 16-16), C02 (or its hydrated ion, HCO ) is activated by attachment to biotin before its transfer to the substrate, in this case the propionate moiety. Formation of the carboxybiotin intermediate requires energy, which is provided by the cleavage of ATP to ADP and Pi- The D-methylmalonyl-CoA thus formed is enzymatically epimerized to its l stereoisomer by methylmalonyl-CoA epimerase (Pig. 17-11). The L-methylmal onyl -CoA then undergoes an intramolecular rearrangement to form succinyl-CoA, which can enter the citric acid cycle. This rearrangement is catalyzed by methylmalonyl-CoA mutase, which requires as its coenzyme 5 -deoxyadenosyl-cobalamin, or coenzyme Bi2, which is derived from vitamin B12 (cobalamin). Box 17—2 describes the role of coenzyme B12 in this remarkable exchange reaction. 

The mechanism of the cleavage of the pyruvate in Eq. 15-37 is not obvious. Thiamin diphosphate is not involved, and free C02 is not formed. The first identified intermediate is an acetyl-enzyme containing a thioester linkage to a cysteine side chain. This is cleaved by reaction with CoA-SH to give the final product. A clue came when it was found by Knappe and coworkers that the active enzyme, which is rapidly inactivated by oxygen, contains a long-lived free radical.326 Under anaerobic conditions cells convert the inactive form E to the active form Ea by an enzymatic reaction with S-adenosylmethionine and reduced flavodoxin Fd(red) as shown in Eq. 15-38.327-329 A deactivase reverses the process.330... 

---