This document discusses dioxygenase enzymes, which activate dioxygen through metal cofactors to incorporate both oxygen atoms into reaction products. It focuses on catechol dioxygenases and arene dioxygenases. Catechol dioxygenases include intradiol and extradiol enzymes that cleave catechol through different mechanisms using Fe(III) and Fe(II), respectively. Model complexes have provided insights into these mechanisms. Arene dioxygenases catalyze cis-dihydroxylation of aromatic rings using non-heme Fe(II). Model complexes have achieved this reaction through proposed catalytic cycles. The document reviews the structures, mechanisms and model chemistries of these important dioxygenase enzyme families.

1. SUDHIR MUDULI (2018MSCH012)
DIOXYGENASE

2. contents
1. Introduction
1.1.Dioxygenase enzyme
1.2.Oxygen activation by metal ions
2.Catechol dioxygenase
2.1. Intradiol catechol dioxygenases
2.2. Model chemistry for intradiol catechol cleavage
2.3. Extradiol catechol dioxygenases
2.4. Model chemistry for extradiol catechol cleavage3
3. Arene (dihydroxylating) dioxygenases
3.1Napthalene dioxygenase: structure and catalytic mechanism
3.2. Model reactions for non-heme iron-catalysed cis-dihydroxylation

3. 1.introduction
• A number of enzymes found in Nature are able to
catalyse the activation of dioxygen from the
atmosphere, and use it to effect a wide variety of
remarkable reactions.
• Enzymes that are able to activate dioxygen are divided
into oxidases(which use oxygen as an oxidant, and reduce
dioxygen to hydrogen peroxide or water) and oxygenases
(which incorporate oxygen atoms from dioxygen into the
product).
Oxygeanse are of two types :mono-oxygenase and
dioxygenase

4. 1.1.dioxygenase enzyme
• Mono-oxygenases catalyse the incorporation of one atom
of oxygen into the product, while di-oxygenases
incorporate both atoms of oxygen into the product(s).
• Most dioxygenase enzymes require a metal cofactor,
which is most often iron(II) or iron(III). Here we will
concentrate on families of nonheme iron(II)-dependent
dioxygenases, for which recent model chemistry is
available. Also we will focus one heme-dependent
dioxygenase, and a small number of cofactor-independent
dioxygenases that have recently been discovered.

5. 1.2. Oxygen activation by metal ions
• Reactions involving the oxidation of hydrocarbons by
dioxygen are exothermic, since the sum of the bond energies
of bonds formed is greater than the sum of the bond energies
of the bonds broken.
• In spite of this exothermicity, dioxygen is chemically
unreactive in the absence of a suitable catalyst.
• The reason is that the ground state for dioxygen (3O2)
contains two unpaired electrons in the highest occupied p*
orbitals, and is therefore spin forbidden to react with spin-
paired singlet species. In contrast, the singlet excited state of
dioxygen (1O2), which contains a pair of valence electrons, is
highly reactive towards alkenes and dienes, as shown in Fig1.

6. 1.2. Oxygen activation by metal ions
• However, since 1O2 is 22 kcal/mol higher in energy than 3O2, it is
not feasible for oxygenase enzymes to access this excited state.
• Transition metal ions containing unpaired electrons can use
three strategies to activate dioxygen.

7. 1.2. Oxygen activation by metal ions
• Orbital overlap with a metal ion: Upon complexation of dioxygen to a
transition metal ion containing unpaired 3d electrons, the unpaired
electrons in the dioxygen p* orbitals are able to overlap with those on the
metal ion.
• Single electron transfer :The transition metals found in metallo-enzymes
that activate dioxygen have two consecutive available oxidation states
(e.g. Fe(II)/Fe(III), Cu(I)/-Cu(II)), hence the metal centre is able to carry out
single electron transfer to bound dioxygen.
• We will see this in extradiol catechol dioxygenases.
• Reaction with a substrate radical : Since the reaction of dioxygen via
radical mechanisms is a spin-allowed process,it is an alternative possible
mechanism.
• It has been proposed that a substrate activation mechanism of this kind
occurs in the intradiol catechol dioxygenases, where a bound catechol
semiquinone intermediate attacks dioxygen to form a hydroperoxide
radical

8. 2. Catechol dioxygenases
• The catechol dioxygenases catalyse the oxidative cleavage of
catechol and substituted catechols, a key step in the bacterial
degradation of aromatic compounds in the environment.
• Two families of dioxygenase enzyme were discovered by Hayaishi
which can catalyse the oxidative cleavage of catechol, both families
utilising dioxygen as a substrate (Fig. 2).
• The intradiol dioxygenases, typified by catechol 1,2-dioxygenase (or
pyrocatechase), cleave the carbon–carbon bond betweenthe phenolic
hydroxyl groups to yield muconic acid as the product, and require
Fe(III) as a cofactor.
• The extradiol dioxygenases, typified by catechol 2,3-dioxygenase
(or metapyrocatechase), cleave the carbon–carbon bond adjacent to
the phenolic hydroxyl groups to yield 2-hydroxymuconaldehyde as
the product, and require Fe(II) as a cofactor.

9. 2. Catechol dioxygenases
• Genereal representation

10. 2. Catechol dioxygenases

11. 2.1. Intradiol catechol dioxygenase
• The first X-ray structure of a catechol dioxygenase, the
intradiol-cleaving protocatechuate 3,4-dioxygenase (3,4-PCD)
from Pseudomonas putida, was solved by Ohlendorf et al.

12. 2.1.Intradiol catechol dioxygenase :
proposed mechanism

13. 2.2. Model chemistry for intradiol
catechol cleavage
The first model system for intradiol cleavage was an Fe(III)–
nitrilotriacetate (NTA)
complex which was reported
to convert 3,5-di-tert-butylca-
techol catalytically over a period
of four days in the presence of
oxygen to give the furanone
derivative in 80% yield.
The FeIII–nitrilotriacetate complex 1 showed the highest
reactivity, and the highest redox potential of þ59 mV (and
hence the highest affinity of thecatechol ligand for the Fe(III)
centre)

14. 2.2. Model chemistry for intradiol
catechol cleavage
• Que and co-workers led to the discovery of more reactive Fe(III)
complexes, the most active of which was Fe(III)–tris(2-
pyridylmethyl)amine (TPA).This complex was found to react with dioxygen
within minutes to form furanone 2 in 98% yield, at a rate of 15 M21 s21,
approximately 1000-fold faster than complex1 Analysis of complex 3 by X-
ray crystallography and 1H NMR spectroscopy revealed a very strong iron–
catecholate interaction, and increased semiquinone character in the
bound substrate

15. 2.2. Model chemistry for intradiol
catechol cleavage
• A catalytically active model system has been reported by Kruger et al.
using N,N0-dimethyl-2,11-diaza[3,3](2,6)pyridinophane (11) as a
macrocyclic ligand.
• Yield =54%
• Time= 30hr

16. Structures of catechols, ligands,
(catecholato)iron(III) complexes, and products
• Structure of catechols
• H2CatA = protochatechuic acid
• H2CatB =3,5ditert-butylcatechol
• H2CatC = pyrochatechol
• H2CatD = 4-chlorocatechol
• H2CatE = 3-chlorocatechol
• H2CatF =2,3,4,5-tetrachlorocatechol
• H2CatG = 3-methoxycatechol
Fig. 3. List of catechols

17. • Structure of ligands
• Tetradentate planar (L2 ,L3) ,tetradentate tripodal (L4 - L24 and L29) tetradentate
linear (L25 - L28), tetradentate tetraazamacrocyclic (L30 , L31),tridentate(L32 – L40)
Fig.4. List of various ligands

18. • By employing these ligands, the structures of 18 (catecholato)iron(III)complexes
have been determined by X-ray analysis.

19. 2.3. Extradiol catechol dioxygenases
• The extradiol catechol dioxygenases catalyse the oxidative
cleavage of the carbon–carbon bond adjacent to the phenolic
hydroxyl groups, to give a 2-hydroxymuconaldehyde
product, using iron(II) as a cofactor.
Catechol 2,3-dioxygenase tetramer

20. 2.3. Extradiol catechol dioxygenases
• proposed mechanism

21. 2.4. Model chemistry for extradiol
catechol cleavage
• Funabiki et al. found that FeCl3 complexes with bipyridine/pyridine
prepared in situ cleave 3,5-di-tert-butylcatechol to give orthoquinone(20),
intradiol products( pyrones 18 and 19).
• Dei et al. obtained a 35% yield using complex [FeIII- (TACN)(dbc)]Cl (21),
while Que et al. used the same complex to give an almost quantitative
yield in CH2Cl2 in the presence of AgOTf.

22. Comparative study
TACN model system

23. 3. Arene (dihydroxylating)
dioxygenases
• The initial step in the bacterial degradation of arene hydrocarbons is
usually a cis-dihydroxylation of the aromatic ring, to give a cis-dihydro
diol. This reaction is catalysed by a family of non-heme iron-dependent
dioxygenases, of which the best studied is naphthalene dioxygenase

24. Proposed catalytic cycle for a model
dihydroxylation reaction

25. 3.2. Model reactions for non-heme
iron-catalysed cisdihydroxylation
• In 1999, Chen and Que reported that an iron (II) complex of 6-trimethyl-
TPA (25) could catalyse the cis-dihydroxylation of cyclooctene, using
H2O2 as oxidant.
• Further tetradentate ligands were subsequently found that could catalyse
this reaction: ligand 26 complexed with iron(II) gave a 75% yield of epoxide
product, while the chiral ligand 27 gave an enantiomeric excess of 82% in
the cis-diol product formed from trans-2-octene

26. Reference
• R. Yamahara et al. : Inorganica Chimica Acta 300–302 (2000) 587–596
• R. Yamahara et al. / Journal of Inorganic Biochemistry 88 (2002) 284 –294
• C.-H. Wang et al. / Inorganica Chimica Acta 360 (2007) 2944–2952
• M. Palaniandavar, R. Mayilmurugan / C. R. Chimie 10 (2007) 366e379
• K. Visvaganesan et al. / Inorganica Chimica Acta 378 (2011) 87–94
• M. Velusamy et al. / Journal of Inorganic Biochemistry 99 (2005) 1032–1042