Ribonucleotide reductase metal-dependent

The function of the metal site in the oxygen-dependent radical enzymes galactose oxidase, amine oxidases, ribonucleotide reductase, and cytochrome c oxidase is inter alia to bind 02 in their reduced forms and undergo the appropriate redox chemistry to generate a metal-bound, activated oxygen species of variable nature. 

The next five transition metals iron, cobalt, nickel, copper and zinc are of undisputed importance in the living world, as we know it. The multiple roles that iron can play will be presented in more detail later in Chapter 13, but we can already point out that, with very few exceptions, iron is essential for almost all living organisms, most probably because of its role in forming the amino acid radicals required for the conversion of ribonucleotides to deoxyribonucleotides in the Fe-dependent ribonucleotide reductases. In those organisms, such as Lactobacilli6, which do not have access to iron, their ribonucleotide reductases use a cobalt-based cofactor, related to vitamin B12. Cobalt is also used in a number of other enzymes, some of which catalyse complex isomerization reactions. Like cobalt, nickel appears to be much more extensively utilized by anaerobic bacteria, in reactions involving chemicals such as CH4, CO and H2, the metabolism of which was important... 

The use of a mixed-valent, dinuclear iron site, similar to those in hemerythrin and ribonucleotide reductase,to catalyze a nonredox reaction such as phosphate ester hydrolysis is novel and unexpected for a variant of the familiar oxo(hydroxo)-bridged diiron center. In contrast to the general agreement that exists regarding the spectroscopic and physical properties of the PAPs, their kinetics properties and especially their mechanism of action remain controversial. Much of the disagreement stems from the different pH dependences of the catalytic activity of BSPAP and Uf, which is due to the fact that the former is isolated in a proteolytically activated form while the latter is not. Proteolysis results in a substantial increase in optimal pH in addition to an increase in catalytic activity at the optimal pH. "" Current data suggest that many of the spectroscopic studies described in the literature were performed on a catalytically inactive form of the enzyme. As a result, the roles of the trivalent and divalent metal ions in catalysis and in particular the identity of the nucleophilic hydroxide that directly attacks the phosphate ester remain unresolved. 

The homolytic cleavage of the Co - C bond of the protein-boimd organo-metallic cofactor AdoCbl (2) is the initial step of the coenzyme Bi2-catalyzed enzymatic reactions. Halpern quoted that adenosyl cobamides can be considered as reversibly functioning sources for organic radicals [119]. A neutral aqueous solution of 2 is remarkably stable with a half-Ufe of 10 s (in the dark at room temperature), but decomposes, mainly with the homolysis of the Co-C bond, at higher temperatures [119,123]. The coenzyme B12-catalyzed enzyme reactions occur with maximal rates of approximately 100 s [173,239]. Rapid formation of Co(ll)corrins occurs only with addition of substrate to a solution of holoenzyme (or of apoenzymes and 2), as demonstrated in most of the known coenzyme Bi2-dependent enzymes, e.g., in methyl-malonyl-CoA mutase [121], glutamate mutase [202] and ribonucleotide reductase [239]. 

It is known that in microorganisms ribonucleotide reductase (RNR) can use either AdoCbl, Fe or Mn as cofactors (Reichard, 1962, 1985 Thelander et al., 1983). The activity of the metal-dependent RNR is inhibited by hydroxyurea. The inhibition of RNR by hydroxyurea was demonstrated in P. coccoides, P. jensenii and P. shermanii. In other strains the inhibition has not been observed and this observation is an additional evidence for the presence of metal-independent, AdoCbl-dependent RNR in these strains. 

The thiosemicarbazones (Structure 62, Figure 6.2) have antitumour activity in their own right [62]. Early mechanistic studies led to the postulate that the mode of action is by inhibition of ribonucleotide reductase [63, 64]. This metal-dependent enzyme is a key intermediate in DNA biosynthesis because it converts ribonucleotides to deoxyribo-nucleotides [65,66]. 

The metal dependence of ribonucleotide reductase is cobalt as vitamin Bi2 (prokaryotes) and iron (eukaryotes), with some evidence for manganese as cofactor [66]. The reader is referred to Ref. [66] for a recent discussion on the mechanism of these interesting enzymes. In essence, the reaction can be described ... 

Metal ions are clearly essential for the ribonucleoside triphosphate reductase isolated from the filamentous cyanophyte, Anabaena 7119, one of the many blue-green algae that depend on deoxyadenosylcobalamin for deoxyribonucleotide synthesis The purified enzyme possesses a molecular weight of 72,000 (estimated by gel filtration) with no subunit structure. It does not reduce ribonucleotides in the absence of divalent cations Ca" " is most effective but Mg" " and Mn" also support enzyme catalysis. Judging from their optimum concentration (5-10 mM) the metal ions are not only necessary to complex the substrate triphosphate but should have an effect on the enzyme protein itself. 

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