Oxidation oxygenases

Protease inhibitors are well-characterized chiral drugs in terms of their mechanism of action. An important new class of protease inhibitors comprises molecules designed to treat HIV infection. In particular, indinavir sulfate (CRIXIVAN, Merck and Co., Inc.) contains five chiral centers that must be of a specific orientation for the molecule to have the desired therapeutic effect. Manufacturing processes for these compounds involving chemical synthesis steps can be quite inefficient, due to yield reduction caused by racemization at each step where a chiral center is formed. A key intermediate in the synthesis of CRIXIVAN is cis-(lS,2R)-l-amino-2-indanol [(-)-CAI], an indene derivative that contributes two chiral centers to indinavir sulfate (Fig. 1). To circumvent the technically demanding chemical synthesis of (-)-CAI and reduce product loss, Merck scientists conceptualized a bioconversion process in which indene is oxidized to one of three derivatives that can serve as precursors to (-)-CAI cis-(lS,2R)-indandiol, trans-(lR,2R)-indandiol, or (lS,2R)-indan oxide. Oxygenases that have been identified in isolates of the genus Pseudomonas and Rhodococcus can catalyze this transformation. 

Nothing is known about the identity of the iron species responsible for dehydrogenation of the substrate. Iron-oxo species such as FeIV=0 or Fem-OOH are postulated as the oxidants in most heme or non-heme iron oxygenases. It has to be considered that any mechanistic model proposed must account not only for the observed stereochemistry but also for the lack of hydroxylation activity and its inability to convert the olefinic substrate. Furthermore, no HppE sequence homo-logue is to be found in protein databases. Further studies should shed more light on the mechanism with which this unique enzyme operates. 

The lower than expected yields can be explained by the nature of methane oxidation to methanol in these bacteria. This reaction, catalysed by methane mono-oxygenase, is a net consumer of reducing equivalents (NADH), which would otherwise be directed to ATP generation and biosynthesis. In simple terms the oxidation of methane to methanol consumes energy, lowering the yield. 

P450 Mono-oxygenase System Reactive Oxygene Species Oxidative Stress Vitamin C Vitamin E... 

All mammalian cells are virtually capable of producing CO with heme as the main substrate (Fig. 1) [5]. Enzymatic heme metabolism in vivo is mainly catalyzed by heme oxygenase (HO). In the presence of HO, the porphyrin ring of heme is broken and oxidized at the a-methene bridge, producing equimolar amounts of CO, ferrous iron, and biliverdin. Three isoforms of HO have been identified. Inducible HO-1 (32 kDa) is mostly recognized for its upregulation in response... 

These include the mitochondrial respiratory chain, key enzymes in fatty acid and amino acid oxidation, and the citric acid cycle. Reoxidation of the reduced flavin in oxygenases and mixed-function oxidases proceeds by way of formation of the flavin radical and flavin hydroperoxide, with the intermediate generation of superoxide and perhydroxyl radicals and hydrogen peroxide. Because of this, flavin oxidases make a significant contribution to the total oxidant stress of the body. 

The second pathway is the eccentric cleavage that occurs at double bonds other than the central 15,15 -double bond of the P-carotene molecule to produce different products called P-apocarotenals with various chain lengths. Because only trace amounts of apocarotenals were detected in vivo from tissues of animals fed P-carotene and these compounds can be formed non-enzymatically from P-carotene auto-oxidation, the existence of this pathway was controversial until recently. The identification of P-carotene 9, 10 -oxygenase (BC02), which acts specifically at the 9, 10 double bond of P-carotene to produce P-apo-lO -carotenal and P-ionone, provided clear evidence of the eccentric cleavage pathway in vivo. Lycopene was also reported as a substrate for BC02 activity. 

Braaz R, P Fischer, D Jendrossek (2004) Novel type of heme-dependent oxygenase catalyzes oxidative cleavage of rubber (poly-cw-l,4-isoprene). Appl Environ Microbiol 70 7388-7395. 

Wackett LP, GA Brusseau, SR Householder, RS Hanson (1989) Survey of microbial oxygenases trichloroethylene degradation by propane-oxidizing bacteria. Appl Environ Microbiol 55 2960-2964. 

Dalton H, BT Golding, BW Waters, R Higgins, JA Taylor (1981) Oxidation of cyclopropane, methylcyclopro-pane, and arenes with the mono-oxygenase systems from Methylococcus capsulatus. J Chem Soc Chem Commun 482-483. 

Lee J, M Simurdiak, H Zhao (2005) Reconstitution and characterization of aminopyrrolnitrin oxygenase, a Rieske iV-oxygenase that catalyzes unusual arylamine oxidation. J Biol Chem 280 36719-36727. 

Sparrow LG, PPK Ho, TK Sundaram, D Zach, EJ Nyns, EE Snell (1969) The bacterial oxidation of vitamin Bg. VII. Purification, properties, and mechanism of action of an oxygenase which cleaves the 3-hydroxypyridine ring. J Biol Chem 244 2590-2660. 

Maeng JH, Y Sakai, Y Tani, N Kato (1996) isolation and characterization of a novel oxygenase that catalyzes the first step of -alkane oxidation in Acinetobacter sp. strain M-1. J Bacteriol 178 3695-3700. 

Latham J, AC Walsh (1986) Retention of configuration in oxidation of a chiral boronic acid by the flavoenzyme cyclohexanone oxygenase. J Chem Soc Chem Commun 527-528. 

Erwin DP, IK Erickson, ME Delwiche, FS Colwell, JL Strap, RL Crawford (2005) Diversity of oxygenase genes from methane- and ammonia-oxidizing bacteria in the Eastern Snake River plain aquifer. Appl... 

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... 

In the rat retina, ischemia upregulates expression of the neuronal nitric oxide synthase and cyclo-oxygenase-2 these effects can be effectively inhibited by lutein (Choi et al., 2006). 

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