Dehydrogenases xanthine dehydrogenase

RUBREDOXIN NAD REDUCTASE SERINE DEHYDROGENASE SORBITOL DEHYDROGENASE XANTHINE DEHYDROGENASE NAD formation,... 

Chemiluminescence and bioluminescence are also used in immunoassays to detect conventional enzyme labels (eg, alkaline phosphatase, P-galactosidase, glucose oxidase, glucose 6-phosphate dehydrogenase, horseradish peroxidase, microperoxidase, xanthine oxidase). The enhanced chemiluminescence assay for horseradish peroxidase (luminol-peroxide-4-iodophenol detection reagent) and various chemiluminescence adamantyl 1,2-dioxetane aryl phosphate substrates, eg, (11) and (15) for alkaline phosphatase labels are in routine use in immunoassay analyzers and in Western blotting kits (261—266). 

Example of a Protein Purification Scheme Purification of the Enzyme Xanthine Dehydrogenase from a Eungus... 

Most purification procedures for a particular protein are developed in an empirical manner, the overriding principle being purification of the protein to a homogeneous state with acceptable yield. Table 5.5 presents a summary of a purification scheme for a selected protein. Note that the specific activity of the protein (the enzyme xanthine dehydrogenase) in the immuno-affinity purified fraction (fraction 5) has been increased 152/0.108, or 1407 times the specific activity in the crude extract (fraction 1). Thus, xanthine dehydrogenase in fraction 5 versus fraction 1 is enriched more than 1400-fold by the purification procedure. 

The enzymes that utilize molybdenum can be grouped into two broad categories (1) the nitrogenases, where Mo is part of a multinu-clear metal center, or (2) the mononuclear molybdenum enzymes, such as xanthine oxidase (XO), dimethyl sulfoxide (DMSO) reductase, formate dehydrogenase (FDH), and sulfite oxidase (SO). The last... 

The three known crystal structures of molybdopterin-containing enzymes are from members of the first two families the aldehyde oxido-reductase from D. gigas (MOP) belongs to the xanthine oxidase family (199, 200), whereas the DMSO reductases from Rhodobacter (R.) cap-sulatus (201) and from/ , sphaeroides (202) and the formate dehydrogenase from E. coli (203) are all members of the second family of enzymes. There is a preliminary report of the X-ray structure for enzymes of the sulfite oxidase family (204). 

Xanthine dehydrogenase that mediates the conversion of hypoxanthine into xanthine and uric acid has been studied extensively since it is readily available from cow s milk. It has also been studied (Leimkiihler et al. 2004) in the anaerobic phototroph Rhodobacter capsulatus, and the crystal structures of both enzymes have been solved. Xanthine dehydrogenase is a complex flavoprotein containing Mo, FAD, and [2Fe-2S] redox centers, and the reactions may be rationalized (Hille and Sprecher 1987) ... 

Hetterich D, B Peschke, B Tshisuaka, F Lingens (1991) Microbial metabolism of quinoline and related compounds. X. The molybdopterin cofactors of quinoline oxidoreductases from Pseudomonas putida 86 and Rhodococcus sp. B1 and of xanthine dehydrogenase from Pseudomonas putida 86. Biol Chem Hoppe-Seyler 372 513-517. 

Koenig K, JR Andreesen (1990) Xanthine dehydrogenase and 2-furoyl-coenzyme A dehydrogenase from Pseudomonasputida Ful two molybdenum-containing dehydrogenases of novel structural composition. J Bacterial 172 5999-6009. 

Leimkiihler S, AL Stockert, K Igarashi, T Nishino, R Hille (2004) The role of active site glutamate residues in catalysis of Rhodobacter capsulatus xanthine dehydrogenase. J Biol Chem 279 40437-40444. 

Self WT (2002) Regulation of purine hydroxylase and xanthine dehydrogenase from Clostridium purinolyti-cum in response to purines, selenium and molybdenum. J Bacterial 184 2039-2044. 

The anaerobe Peptococcus (Micrococcus) aerogenes had a dehydrogenase that carried out specific hydroxylation at the 6-positions of 2- and 8-hydroxypurine, and was therefore distinct from xanthine dehydrogenase from which it could be separated (Woolfolk et al. 1970). It was also able to carry out dismutation of 2-hydroxypurine to xanthine (2,6-dihydroxypurine) and hypoxanthine (6-hydroxypurine). 

Although it had been assumed that only hypoxanthine dehydrogenase is required for the conversion of hypoxanthine (6-hydroxypurine) into uric acid, in Clostridium purinolyti-cum, two enzymes, both of which contain a selenium cofactor, are required. The enzymes differ in the molecular mass of their subunits, in their terminal amino acid sequences, in their kinetic parameters, and in their specific activities for purines (Self and Stadman 2000). Purine hydroxylase converts purine into hypoxanthine and xanthine (2,6-dihy-droxypurine), which is then further hydroxylated to uric acid (2,6,8-trihydroxypurine) by xanthine dehydrogenase (Self 2002). 

Self WT, TC Stadman (2000) Selenium-dependent metabolism of purines a selenium-dependent purine hydroxylase and xanthine dehydrogenase were purified from Clostridium purinolyticum and characterized. Proc Natl Acad Sci USA 97 7208-7213. 

Wagner R, R Cammack, JR Andreesen (1984) Purification and characterization of xanthine dehydrogenase from Clostridium acidiurici grown in the presence of selenium. Biochim Biophys Acta 791 63-74. 

Xi H, BL Schneider, L Reitzer (2000) Purine catabolism in Escherichia coli and function of xanthine dehydrogenase in purine salvage J Bacterial 182 5332-5341. 

Increased levels of cytosolic calcium could potentiate ischaemia-reperfusion injury in several ways. For example, conversion of xanthine dehydrogenase to xanthine oxidase may be catalysed by a calcium-dependent protease (McCord, 1985). However, because it has been so difficult to demonstrate the presence of xanthine... 

Roy, R.S. and McCord, J.M. (1983). Superoxide and ischaemia conversion of xanthine dehydrogenase to xanthine oxidase. In Oxyradicals and their Scavenging Systems , Vol. 2 (eds. K Greenwald and G. Cohen) pp. 145-153. Elsevier, New York. 

Immunohistochemical studies carried out in our laboratories have demonstrated the presence of xanthine oxidase in synovial endothelial cells (Stevens etal., 1991). As expected, the activity of this enzyme per unit weight of tissue is generally higher in synovia taken from RA patients due to their increased vascularity (Allen et al., 1987). In addition, it has also been shown that rheumatoid synoviocytes contain increased levels of iron-saturated ferritin (Morris et d., 1986). Xanthine oxidase (but not dehydrogenase) is able to mobilize iron from ferritin, supplying the necessary transition metal catalyst for the Haber-Weiss reaction and promoting OH formation (Biemond eta/., 1986). 

Reiners, J.J. and Rupp, T. (1989). Conversion of xanthine dehydrogenase to xanthine oxidase occurs during keratinocyte differentiation modulation by 12-O-tetradecanoylphorbol-13-acetate. J. Invest. Dermatol. 93, 132—135. 

Stirpe, F. and Della Corte, E. (1969). The regulation of rat liver xanthine oxidase. Conversion in vitro of the enzyme activity from dehydrogenase (type D) to oxidase (type O). J. Biol.Chem. 244, 3855-3863. 

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