Xylene monooxygenases

Some of the major enzyme groups that facilitate this transformation are heme-containing MOs of the cytochrome P450 type [111], alkane hydroxylases, xylene monooxygenases, styrene monooxygenases [105], and haloperoxidases [112],... 

Styrene was successfully oxidized to the S-product both by xylene monooxygenase from P. putida mt-2 [113] and styrene monooxygenase from Pseudomonas sp.VLB120 [114] (Scheme 9.13), with the latter enzyme displaying a particularly large substrate tolerance with excellent stereoselectivity (>99% ee). In this context it is interesting to note that both xylene monooxygenase as well as chloroperoxidase are very selective for mono-epoxidation in case of presence of multiple alkene functionalities [115]. 

Toluene/o-xylene monooxygenase in P. stutzeri strain 0X1 carried ont snccessive monooxygenation of o-xylene (Bertoni et al. 1998), and the tolnene-4-monooxygenase of Pseudomonas mendocina KRl and tolnene-3-monooxygenase of Ralstonia pickettii PKOl can hydroxylate benzene, tolnene, and o-xylene (Tao et al. 2004 Vardar and Wood 2004). 

Bertoni G, M Martino, E Galli, P Barbieri (1998) Analysis of the gene cluster encoding toluene/o-xylene monooxygenase from Pseudomonas stutzeri 0X1. Appl Environ Microbiol 64 3626-3632. 

Suzuki M, T Hayakawa, JP Shaw, M Rekik, S Harayama (1991) Primary structure of xylene monooxygenase similarities to and differences from the alkane hydroxylation system. J Bacteriol 173 1690-1695. 

Vardar G, TK Wood (2004) Protein engineering of toluene-o-xylene monooxygenase from Pseudomonas stutzeri 0X1 for synthesizing 4-methylresorcinol, methylhydroquinone, and pyrogallol. Appl Environ Microbiol 70 3253-3262. 

The pathway for the degradation of the xylenes depends critically on the orientation of the methyl groups, and o-xylene is considered to be the most recalcitrant since xylene monooxygenase cannot hydroxylate one of its methyl groups ... 

Fe—2S] ferredoxins, 38 238-239 xylene monooxygenase, 38 239 Pseudorotation, square-planar substitution reactions, 34 239-247 5-coordinate intermediate, 34 245... 

Shanklin J., Whittle E. and Fox B. G. (1994) Eight histidine residues are catalytically essential in a membrane associated iron enzyme, stearoyl-CoA desaturase and are conserved in alkane hydroxylase and xylene monooxygenase. Biochem. 33, 12787-12794. 

Figure 1. Classification of oxidoreductases with a focus on enzymes capable of sp3-carbon oxyfunctionalization. MMO, methane monooxygenase AMO, alkane monooxygenase XMO, xylene monooxygenase. The most prominent enzyme classes in respect of sp3-carbon oxyfunctionalization are shown in bold.

Ryoo D, Shim H, Canada K, Barbieri P, Wood TK (2000) Aerobic degradation of tetra-chloroethylene by toluene-o-xylene monooxygenase of Pseudomonas stutzeri OX1. Nat Biotechnol 18 775-778... 

Austin, R.N., K. Buzzi, E. Kim, G.B. Zylstra, and J.T. Groves (2003). Xylene monooxygenase, a membrane-spanning non-heme diiron enzyme that hydroxylates hydrocarbons via a substrate radical intermediate. J. Biol. Inorg. Chem. 8, 733-739. 

ACP = acyl carrier protein ACPA D = ACPA desat-urase AlkB = octane 1-monooxygenase AOX = alternative oxidase DMQ hydroxylase = 5-demethoxyquinone hydroxylase EXAFS = extended X-ray absorption fine structure spectroscopy FMN = flavin mononucleotide FprA = flavoprotein A (flavo-diiron enzyme homologue) Hr = hemerythrin MCD = magnetic circular dichroism MME hydroxylase = Mg-protophorphyrin IX monomethyl ester hydroxylase MMO = methane monooxygenase MMOH = hydroxylase component of MMO NADH = reduced nicotinamide adenine dinucleotide PAPs = purple acid phosphatases PCET = proton-coupled electron transfer, PTOX = plastid terminal oxidase R2 = ribonucleotide reductase R2 subunit Rbr = rubrerythrin RFQ = rapid freeze-quench RNR = ribonucleotide reductase ROO = rubredoxin oxygen oxidoreductase XylM = xylene monooxygenase. 

A very convenient green synthesis of epoxystyrene has been developed. It consist of the reaction of styrene with xylene monooxygenase, which introduces an epoxide in the vinyl double bond of styrene (Scheme 26). 

E. coli recombinants carrying only xylene monooxygenase system... 

Scheme 3.11 Oxidation of toluene derivatives toward benzoic acid analogs (R = H, CHj) catalyzed by a gene cluster containing xylene-monooxygenase (XMO), benzylalcohol dehydrogenase (BADH), and benzaldehyde dehydrogenase (BZDH).

Buehler, B., Schmid, A., Hauer, B., and Witholt, B. (2000) Xylene monooxygenase catalyzes the multistep oxygenation of toluene and pseudocumene to corresponding alcohols, aldehydes, and acids in Escherichia coli JMIOI. J. Biol Chem., 275, 10085-10092. 

Figure 12. Active site structure of toluene/o-xylene monooxygenase (ToMOH) from Pseudomonas stutzeri 0X1 (A) fully oxidized native ToMOH (PDB code 2INC), (B) Mn /Mn ToMOH (PDB code 2IND), and (C) reduced ToMOH-azide complex (PDB code ITOR).

Figure 13. EPR spectra of the Fe -Fe" -W intermediate generated in the reaction of reduced mow mutant of toluene/o-xylene monooxygenase with dioxygen. Reprinted with permission from [214]. Copyright 2006, American Chemical Society.

Cafaro V, Scognamiglio R, Viggiani A, Izzo V, Passaro I, Notomista E, Dal Piaz F, Amoresano A, Casbarra A, Pucci P, Di Donato A. 2002. Expression and purification of the recombinant subunits of toluene/o-xylene monooxygenase and reconstitution of the active complex. Eur JBiochem 269 5689-5699. 

Sazinsky MH, Bard J, Di Donato A, Lippard SJ. 2004. Crystal structure of the toluene/o-xylene monooxygenase hydroxylase from Pseudomonas stutzeri 0X1 insight into the substrate specificity, substrate channeling, and active site tuning of multicomponent monooxygenases. JBiol Chem 279 30600-30610. 

Rudd DJ, Sazinsky MH, Lippard SJ, Hedman B, Hodgson KO. 2005. X-ray, absorption spectroscopic study of the reduced hydroxylases of methane monooxygenase and toluene/o-xylene monooxygenase differences in active site structure and effects of the coupling proteins MMOB and ToMOD. Inorg Chem 44 4546-4554. 

McCormick MS, Sazinsky MH, Condon KL, Lippard SJ. 2006. X-ray crystal structures of manganese(II)-reconstituted and native toluene/o-xylene monooxygenase hydroxylase reveal rotamer shifts in conserved residues and an enhanced view of the protein interior. J Am Chem Soc 128 15108-15110. 

Murray LJ, Garcia-Serres R, Naik S, Huynh BH, Lippard SJ. 2006. Dioxygen activation at non-heme diiron centers characterization of intermediates in a mutant form of toluene/o-xylene monooxygenase hydroxylase. JAm Chem Soc 128 7458-7459. 

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