FAD synthase

This enzyme [EC 2.7.T.2], also known as FAD synthase and FAD pyrophosphorylase, catalyzes the ATP-depen-dent transfer of an adenylyl group to FMN to yield FAD and diphosphate. 

The conversion of riboflavin to flavin mononucleotide (FMN) is catalyzed by flavokinase (Figure 9.73). This conversion may occur during absorption through the gut mucosa or in other organs. The subsequent conversion of FMN to flavin adenine dinucleotide (FAD) is catalyzed by FAD synthase. FAD synthase uses ATP as a source of an adenylyl group, in this conversion (McCormick et ah, 1997). Various phosphatases, including those of the gut mucosa, can catalyze the breakdown of FAD to FMN and of FMN to free riboflavin. Dietary flavins that are covalently botmd to proteins are thought to be unavailable and not to contribute to our dietary needs (Bates et ah, 1997). 

It is still unknown how the pyrimidine intermediate 5 is dephosphorylated (reaction VI). However, it is well established that the dephosphorylation product 6 is condensed with 3,4-dihydroxy-2-butanone 4-phosphate (8) by the catalytic action of lumazine synthase (reaction VIII). The carbohydrate substrate 8 is in turn obtained from ribulose phosphate (7) by a complex reaction sequence that is catalyzed by a single enzyme, 3,4-dihydroxy-2-butanone 4-phosphate synthase (reaction VII). As mentioned above, the lumazine 9 is converted to riboflavin (10) by the catalytic action of riboflavin synthase (reaction IX). Ultimately, riboflavin is converted to the coenzymes, riboflavin 5 -phosphate (flavin mononucleotide (FMN), 11) and flavin adenine dinucleotide (FAD, 12) by the catalytic action of riboflavin kinase (reaction X) and FAD synthase (reaction XI). These reaction steps are required in all organisms, irrespective of their acquisition of riboflavin from nutritional sources or by endogenous biosynthesis. 

FIGURE 32-7 Sources of free radical formation which may contribute to injury during ischemia-reperfusion. Nitric oxide synthase, the mitochondrial electron-transport chain and metabolism of arachidonic acid are among the likely contributors. CaM, calcium/calmodulin FAD, flavin adenine dinucleotide FMN, flavin mononucleotide HtT, tetrahydrobiopterin HETES, hydroxyeicosatetraenoic acids L, lipid alkoxyl radical LOO, lipid peroxyl radical NO, nitric oxide 0 "2, superoxide radical. 

NO synthase with cofactors NADPH2 heme, Ca + calmodulin, tetrahydrobiopterin, FAD, FMN... 

Prosthetic groups contained within NOS. Nitric oxide synthases are isolated containing approximately one molecule each of heme, FAD, and FMN per subunit, and also contain variable quantities of tetrahydrobiopterin (0.1 to 1 molecule per subunit). 

Studies on partially purified preparations of HC NO synthase indicated a similar dependency on tetrahydrobiopterin (BH4), reduced glutathione (GSH), FAD, and NADPH as described for the macrophage (Kwon et al., 1989 Stuehr et al., 1989b, Tayeh and Marietta, 1989) (see Table 1). Similar data have been obtained using cytosol from human HC stimulated with cytokines + LPS in culture. 

Figure 3 Schematic representation of nitric oxide synthase isoforms and cytochrome P450 reductase. Haem, heme PDZ, PDZ domain (GLGF repeats) CaM, calmodulin FMN, flavin mononucleotide FAD, flavin adenine dinucleotide (adapted from Hobbs et al., 1999).

Nitric oxide is synthesized from arginine in an NADPH-dependent reaction catalyzed by nitric oxide synthase (Fig. 22-31), a dimeric enzyme structurally related to NADPH cytochrome P-450 reductase (see Box 21-1). The reaction is a five-electron oxidation. Each subunit of the enzyme contains one bound molecule of each of four different cofactors FMN, FAD, tetrahydro-... 

Synthesis of NO Arginine, 02, and NADPH are substrates for cytosolic NO synthase (Figure 13.9). Flavin mononucleotide (FMN), flavin adenine dinucleotide (FAD), heme, and tetrahydro- biopterin are coenzymes for the enzyme, and NO and citrulline are products of the reaction. Three NO synthases have been identified. Two are constitutive (synthesized at a constant rate regardless of physiologic demand), Ca2+-calmodulin-dependent enzymes. They are found primarily in endothelium (eNOS), and neural tissue... 

The first step in valine biosynthesis is a condensation between pyruvate and active acetaldehyde (probably hy-droxyethyl thiamine pyrophosphate) to yield a-acetolactate. The enzyme acetohydroxy acid synthase usually has a requirement for FAD, which, in contrast to most flavopro-teins, is rather loosely bound to the protein. The very same enzyme transfers the acetaldehyde group to a-ketobutyrate to yield a-aceto-a-hydroxybutyrate, an isoleucine precursor. Unlike pyruvate, the a-ketobutyrate is not a key intermediate of the central metabolic routes rather it is produced for a highly specific purpose by the action of a deaminase on L-threonine as shown in figure 21.10. 

Hydroxypropionate / malyl-CoA cycle 10 7 NAD(P)H, but 1 FAD is reduced in the cycle Acetyl-CoA/propionyl-CoA carboxylase HCOJ Acetyl-CoA, pyruvate, succinyl-CoA Malonyl-CoA reductase, propionyl-CoA synthase, malyl-CoA lyase... 

The inherent substrate specificities of Pseudomonas sp. 61-3 PHA synthases (PhaClPs and PhaC2Ps) are rather low toward 3HB monomer (17,18), and the fad mutant E. coli strain used in our study cannot generate enough 3HB monomers from the (3-oxidation pathway (21,26). Therefore, we introduced the R. eutropha phaABRe genes to generate enough more 3HB... 

Figure 21-2. Metabolism of homocysteine. BHMT, betaineihomocysteine methyl-transferase CBS, cystathionine P-synthase Cob, cobalamin CTH, cystathionine y-lyase DHF, dihydrofolate DMG, dimethylglycine FAD, flavin adenine dinucleotide MAT, methionine adenosyltransferase 5-MTHF, 5-methyltetrahydrofolate 5,10-MTHF, 5,10-methylenetetrahydrofolate MTHFR, methylenetetrahydrofolate reductase MS, methionine synthase MTRR, methionine synthase reductase MTs, methyl transferases PLE pyridoxal phosphate SAH, S-adenosylhomocysteine SAHH, SAH hydrolase SAM, 5-adenosylmethionine SHMT, serine hydroxymethyltransferase THF, tetrahydrofolate Zn, zinc.

---