Journal of Bioremediation & Biodegradation

- Open Access

Review Article
OPEN ACCESS Freely available online

www.omicsonline.org

doi:10.4172/2155-6199.1000112

Application of Monooxygenases in Dehalogenation,

Desulphurization, Denitrification and Hydroxylation of
Aromatic Compounds
Pankaj Kumar Arora1, Alok Srivastava2* and Vijay Pal Singh2
1
2

Environmental Biotechnology, Microbial Type Culture Collection and Gene Bank (MTCC), Institute of Microbial Technology, Sector-39A, Chandigarh-160036, India
Department of Plant Science, Faculty of Applied Sciences, M.J.P. Rohilkhand University, Bareilly -243006, India

Abstract
Monooxygenases act as biocatalysts in bioremediation process and synthetic chemistry due to their highly
regioselectivity and sterioselectivity on wide range of substrates. They are involved in the process of desulfurization,
dehalogenation, denitrification, ammonification, hydroxylation, biotransformation and biodegradation of various aromatic
and aliphatic compounds. In the recent years, the practical applications of monooxygenases have been improved using
the approaches of directed evolution, meta-genomics and bioinformatics. This review is focused on current applications
of monooxygenses especially in biodegradation and biotransformation of aromatic compounds.

Keywords: Monooxygenases; Desulfurization; Dehalogenation;

Aromatic compounds; Biodegradation

Introduction
Aromatic compounds are persistence environmental pollutants
that are widely distributed in the biosphere due to anthropogenic
activities. These compounds are highly toxic to living beings; therefore,
many of them have been listed as priority pollutants by United
States Environmental Protection Agency. Microbial degradation has
emerged as an effective technology to remove these compounds
from environment. Many microbial enzymes such as oxygenases,
dehalogenases, reductases, hydroxylases and dehydrogenases are
involved in the degradation of these environmental pollutants. Among
them, oxygenases are the key enzymes for aerobic biodegradation
of aromatic compounds because they are involved in the initial
reaction of degradation and also catalyze the ring cleavage of the
aromatic compounds which is the essential step for the complete
mineralization of these compounds [1].

monooxygenase (TcmH) from Streptomyces glauscens [5]; ActVAorf6 monooxygenase from Streptomycescoelicolor A3(2) [6]; quinol
monooxygenase (YgiN) from E. Coli [7] and Rv0793 a hypothetical
monooxygenase from Mycobacteriumtuberculosis [8].
Aromatic dioxygenase are also divided into two subclasses
based on mode of action: aromatic ring hydroxylation dioxygenases
(ARHDs) and aromatic ring cleavage dioxygenases (ARCDs) [1]. ARHD
catalyzes the incorporation of both atoms of oxygen into aromatic
ring (Figure 1b) whereas ARCD catalyzes ring cleavage of the aromatic

Oxygenases catalyze insertion of one or two oxygen atoms into

the substrates. Two classes of oxygeases have been identified based
on number of oxygen atom used in the oxidation: monooxygenases
and dioxygenases [1]. Monooxygenases incorporate one atom of the
oxygen into the substrate (Figure 1a) whereas dioxygenases add both
atoms of the oxygen [1].

*Corresponding author: Dr. Alok srivastava, Department of Plant Science,

Faculty of Applied Sciences, Rohilkhand University, Bareilly-243006, India, Tel:
+919760821347; E-mail: alok.plsc@rediffmail.com
ReceivedOctober 25, 2010; Accepted November 25, 2010; Published November
27, 2010
Citation: Arora PK, Srivastava A, Singh VP (2010) Application of Monooxygenases
in Dehalogenation, Desulphurization, Denitrification and Hydroxylation of Aromatic
Compounds. J Bioremed Biodegrad 1:112. doi:10.4172/2155-6199.1000112
Copyright: 2010 Arora PK, et al. This is an open-access article distributed under
the terms of the Creative Commons Attribution License, which permits unrestricted
use, distribution, and reproduction in any medium, provided the original author and
source are credited.

Volume 1 Issue 31000112

lishing
Pub
Gr
CS

p
ou

J Bioremed Biodegrad
ISSN:2155-6199 JBRBD, an open access journal

Figure 1: Reactions catalyzed by oxygenases. (a) Monooxygenation of phenol;

(b) aromatic ring hydroxylation of benzene; (c) intradiol aromatic ring cleavage
of catechol; (d) extradiol aromatic ring cleavge of catechol.

OM
I

Monooxygenases are classified into two subclasses based on

which cofactor is present: Flavin dependent monooxygenases and
P450 monooxygenases. Flavin dependent monooxygenases contain
flavin as prostethic group and require NADP or NADPH as coenzyme
[2]. P450 monooxygenases are hame containing oxygenases and
found in both eukaryotic and prokaryrotic organisms. Best example of
bacterial P450 monooxygenase is CYP102 from Bacillusmegaterium
BM3 that is able to hydroxylate a variety of alkanes, fatty acids and
aromatic compounds [3]. Other types of bacterial monooxygenases
are also known that do not contain flavin or hame as a cofactor
e.g., pterin-dependent monooxygenases and metal ion-dependent
monooxygenases [4]. Majority of monooxygenases require cofactor
for their activities, however, some monooxygenases have also been
identified and characterized that do not require any cofactor for their
activities. These cofactor independent monooxygenases require only
molecular oxygen for their activities and utilize the substrate as reducing
agent. Examples of these mononoxygenases are tetracenomycin F1

Citation: Arora PK, Srivastava A, Singh VP (2010) Application of Monooxygenases in Dehalogenation, Desulphurization, Denitrification and
Hydroxylation of Aromatic Compounds. J Bioremed Biodegrad 1:112. doi:10.4172/2155-6199.1000112

Page 2 of 8

Oxygenolytic dehalogenation is further divided into two

classes: dioxygenase type dehalogenation and monooxygenase
type dehalogenation. Dioxygenase type dehalogenase adds two
atoms of oxygen into the substrate to remove the chlorine atom.
They involved in dehalogenation of monochlorinated aromatic
compounds such as 4-chlorophenyl acetate and 2-chlorobenzoate
[14]. Monooxygenase type dehalogenase adds one atom of oxygen
to chlorinated compounds to remove the chlorine atom. Examples
of monooxygenase type dehalogenases are pentachlorophenol
4-monooxygenase, chlorophenol 4-monooxygenase and 2, 4,
6-trichlorophenol monooxygenase. These monooxygenases are
involved in dehalogenation of polychlorinated aromatic compounds
and have been characterized and purified from a variety of
microorganisms.

Pentachlorophenol 4-monooxygenase (EC 1.14.13.50)

ring typically carrying two or more hydroxyl groups [1]. ARCDs are
further divided into two groups based on the ring cleavage: intradiol
(IARCD) and extradiol (EARCD) [1]. Intradiol cleaves the aromatic ring
between two hydroxyl groups (ortho-cleavage) (Figure 1c), whereas
extradiol cleaves the aromatic ring between a hydroxylated carbon
and an adjacent non-hydroxylated carbon (meta-cleavage) (Figure 1d).
Several publications have been focused on dioxygenases and
their role in biodegradation [9-12]. But very few reviews are available
that cover the role of the monooxygenases in biodegradation
[13]. In this review, we have discussed the current applications of
the monooxygenases in biodegradation and biotransformation of
aromatic compounds.

Monooxygenases and Dehalogenation of Polychlorinated

Aromatic Compounds
Polychlorinated compounds are widely distributed in the
environment and cause serious health problem to humans beings
and animals. These compounds are considered recalcitrant due
to presence of multiple chlorine atoms at benzene ring. Examples
of these compounds are polychlorinated biphenyls (PCB),
pentachlorophenol (PCP), dichlorophenol (DCP), trichlorophenol
(TCP), dioxin, 2, 4-dichlorophenoxyacetic acid (2,4-D) and
dichloropheyltrichloroethane (DDT).
Microbial degradation of polychlorinated aromatic compounds
initiated with the removal of the chloride atom from a benzene ring.
Chloride atom can be removed from aromatic ring in three different
ways: hydrolytic, reductive and oxygen dependent dehalogenation
[14]. Hydrolytic dehalogenation includes replacement of chlorine
atom with hydroxyl group. This hydroxyl group is derived from water.
Reductive dehalogenation involves replacement of chlorine atom
with hydrogen atom whereas oxygenolytic dehalogenation involves
replacement of chlorine atom by hydroxyl group whose oxygen atom
is derived from O2.

Chlorophenol 4-monooxygenase (EC 1.14.13.-)

This enzyme is involved in the degradation pathway of 2, 4,
5-trichlorophenoxy acetic acid by Burkholderia cepacia AC1100 [27].
This enzyme catalyzes conversion of 2,4,5-trichlorophenol (2,4,5TCP) to 2,5-dichloro-p-benzoquinone, which is chemically reduced to
Volume 1 Issue 31000112

lishing
Pub
Gr
CS

p
ou

J Bioremed Biodegrad
ISSN:2155-6199 JBRBD, an open access journal

Pentachlorophenol 4-monoxygenase (PcpB) has been purified

and characterized from Sphingomonas chlorophenolica ATCC 39723.
PcpB was comprised of 538 amino acids with molecular weight of
39 kD. PcpB was synthesized in cytoplasm and then translocated to
periplasm via inner memberane [25]. The crystal structure of PcpB
is yet not solved. However, model of 3D structure of PcpB has been
proposed on the basis of homology modelling [26]. The active site
residue has been determined from proposed 3D structure and site
directed mutagenesis was carried out to change the active site residue
[26]. Mutant thus obtained produced mutant proteins that purified
by affinity chromatography. The finding of mutation study supported
the validity of proposed 3D structure based on homology modelling
[26] . On the basis of experiment of site directed mutagenesis of
active site of PcpB, Nakamura et al. (2004) concluded that Phe 85,
Try 216 and Arg 235 are for enzyme activity and Tyr 397 and Phe 87
stablize the structure of the protein [26].

OM
I

Figure 2: Dehalogenations of polychlorinated nitroaromatic compounds by

monooxygenases. (a) Dehalogenation of pentachlorophenol by pentachlorophenol
4-monooxygenase; (b) dehalogenation of 2,4,5-trichlorophenol by chlorophenol
4-monooxygenase; (c) dehalogenation of 2, 4, 6-trichlorophenol by 2, 4,
6-trichlorophenol monooxygenase (d) sequential dehalogenation of 2, 4,
6-trichlorophenol by 2, 4, 6-trichlorophenol monooxygenase

Pentachlorophenol 4-monooxygenase (PcpB) is a flavin

monooxygenase that catalyzes hydroxylation at para position with
removal of the chloride ion in the initial step of the microbial
degradation of pentachlorophenol (Figure 2a). As a result of
hydroxylation, pentachlorophenol converted to tetrahydroquinone
[15]. PcpB also catalyzes reaction on various substituted aromatic
compounds with release of nitro, amino or cyano group from para
position [16]. PcpB is encoded by the pcpB gene that has been
identified in a variety of PCP degrading bacteria [17,18]. Examples
of PCP degrading bacteria are Sphingonium, Sphingomonads and
Pseudomonas. The pcpB gene was identified in the four strains of
Sphingonium chlorophenolica (ATCC 39723, RP-2, SR-3 and ATCC
33790) in which three strains (ATCC 39723, RP-2, SR-3) had identical
pcpB gene sequence [19, 20] whereas the sequence of the pcpB
gene of strain ATCC 33790 was differed by 10% from rest of three
strains [20]. The pcpB gene sequence of Sphinogomonads strain
UG-30 showed 90% sequence similarity with that of Sphingonium
chlorophenolica ATCC 39723 [21-23]. The homologus of pcpB
gene has been identified in polychlorinated degrading bacterium
Novosphingonium sp. strain MT1 [24] as well as in two non-PCP
degrading - and - proteobacterial strains isolated from soil samples
from a PCP-contaminated wood treatment site [18].

Citation: Arora PK, Srivastava A, Singh VP (2010) Application of Monooxygenases in Dehalogenation, Desulphurization, Denitrification and
Hydroxylation of Aromatic Compounds. J Bioremed Biodegrad 1:112. doi:10.4172/2155-6199.1000112

Page 3 of 8

2,5-dichloro-p-hydroquinone (2,5-DiCHQ) (Figure 2b) [28]. The gene

encoding chlorophenol monooxygenase (tftd) has been identified,
sequenced and cloned from Burkholderia cepacia AC1100 and
transformed into E. coli for overexpression [27-30]. The molecular
weight of the purified protein was 58 kD [28]. This protein utilizes
O2, FAD and NADH for catalyzing the reaction but does not utilize
riboflavin and NADPH [28].

2, 4, 6-Trichlorophenol Monooxygenase (EC 1.14.13.-)

This enzyme was first reported in Azotobacter sp. GP1 and catalyzes
conversion of 2,4,6 trichlorophenol to 2,6-dichlorohydroquinone
with release of chloride ion (Figure 2c) [31]. TCP monooxygenase from
Azotobacter was a homotetrameric protein with molecular weight of
240 kD. Another 2,4,6-trichlorophenol (2,4,6-TCP) 4-monooxygenase
was found in Ralstonia eutropha JMP134 that catalyzes sequential
dechlorinations of 2,4,6-TCP to 6-chlorohydroxyquinol [32]. This
monooxygenase oxidize 2, 4, 6-TCP to 2, 6-dichloroquinone that
further hydrolyzed to 2-chlorohydroxyquinone by the same enzyme
(Figure 2d). 2-Chlorohydroxyquinone was then reduced by ascorbate
and NADH to 6-chlorohydroxyquinol [32].

optimum temperature for activity of DszA and DszC was determined

44C and 65C respectively.
Recombinant bacteria have also been created to enhance
the activity of biodesulfurization [48,49]. The recombinant strain
desulfurizes DBT more efficiently than the native one, and also provides
new alternative for the development of a commercial desulfurization
process [50]. A genetically modified organism Pseudomonas putida
CECT 5279 carrying dszABC genes from Rhodococcus erythropolis
IGTS8 and a flavin oxidoreductase (hpac) from E.coli was constructed
for biodesulfurization process [51]. This recombinant strain produced
maximum amount of DszB enzyme at the early exponential phase
that rapidly decreased at the middle exponential phase, hampering
an efficient transformation of HBPS into HBP. To overcome this
problem, a model two-step BDS resting cell process using P. putida
cells from the late and early exponential growth phases was designed
to significantly increase biodesulfurization. In the first step, the cells
of late exponential phase (10 h grown cells) of P. putida CECT 5279
showing the maximum activities of DszA and DszC monooxygenases,

Monooxygenases Involved in Biodesulfurization of

Dibenzothiophene

Although the biochemistry, genetics and physiology of

desulphurization have been extensively studied in Rhodococcus
sp., the desulfurizing genes (dszABC) have also been identified in
other mesophilc bacteria [41-43] as well as thermophilc bacteria.
The sequences of the dszABC genes of a thermophilic bacterium
Mycobacterium phlei that carry out biodesulfurization at range of 2050C, was found to be identical to that of Rhodococcus erythropolis
IGTS8 [44-46]. The DszA and DszC from the thermophilic bacterium
Paenibacillus sp. strain A11-2 have been purified and characterized
[47]. DszA is dimeric protein with molecular weight of 120 kD and
subunit molecular weight of 48 kD whereas DszC was tetrameric
protein with molecular weight of 200 kD and subunit mass 43 kD. The

Figure 4: Monooxygenases involved in denitrification. (a) Conversion of

4-nitrophenol to p-benzaquinone by 4-nitrophenol 4-monooxygenase; (b)
conversion of 4-nitrophenol to 4-nitrocatechol by 4-nitrophenol 2-monooxygenase;
and (c) conversion of 2-nitrophenol to catechol by 2-nitrophenol 2-monooxygenase.

Volume 1 Issue 31000112

lishing
Pub
Gr
CS

p
ou

J Bioremed Biodegrad
ISSN:2155-6199 JBRBD, an open access journal

Figure 3: Biodesulfurization of DBT. (a) conversion of DBT to DBT sulphoxide

by DBT monooxygenase; (b) conversion DBT sulphoxide to DBT sulphone
by DBT monooxygenase; (c) conversion of DBT sulphone to HBPS by DBT
sulfone monooxygenase and (d) conversion of HBPS to HBP by HBPS
desulfinase.

OM
I

Sulphur containing organic compounds such as dibenzothiophene

(DBT) are present in the fossil fuels. On combustion of these fossil
fuels, sulphur dioxide is released into the environment and causes air
pollution. The complete removal of sulphur from these compounds
is not possible by conventional physical and chemical methods.
Biodesulfurization, therefore, is a process that completely removes
sulphur from these organic compounds. Biodesulfurization of DBT is
well characterized in Rhodococcus erythropolis IGTS8 [33]. Two flavin
dependent monooxygenases, DBT monooxygenase (DszC) and DBT
sulfone monooxygenases (DszA) are involved in the initial steps of
the biodesulfurization. DBT monooxygenase (EC 1.14.13.-) converts
DBT to DBT sulphoxide which is further converted to DBT sulphone
by same enzyme (Figure 3a and 3b). DBT sulfone monooxygenase
(EC 1.14.13.-) converts DBT sulfone to 2-hydroxybiphenyl 2-sulfinate
(HBPS) (Figure 3c). Both of the monooxygenases require another
enzyme flavin reductase (DszD) for their activities. One more
enzyme HBPS desulfinase (DszB) is also involved in the final step of
biodesulfurization and converts HBPS to hydroxybiphenyl (HBP) and
sulphate (Figure 3d). In R. erythropolis, enzymes DszA, DszB and DszC
are encoded by plasmid located dsz operon and another enzyme DszD
is considered as genome encoded [34, 35]. The desulfurization genes
was conserved among Rhodococcus species [36]. Several strains of R.
erythropolis, i.e., SY1, D-1, Ni-36, Ni-43, and QIA-22 exhibited same
DBT-desulfurizing reaction as reported in Rhodococcus erythropolis
IGTS8 [37-40].

Citation: Arora PK, Srivastava A, Singh VP (2010) Application of Monooxygenases in Dehalogenation, Desulphurization, Denitrification and
Hydroxylation of Aromatic Compounds. J Bioremed Biodegrad 1:112. doi:10.4172/2155-6199.1000112

Page 4 of 8

were transformed all DBT into 50% mixture of HBPS and HBP [51]. In
the second step, the cells of late exponential phase have removed
and the early exponential phase cells (5 h grown cells) were added.
After addition remaining HBPS was efficiently converted to HBP [51].
Another recombinant bacteria has been design by transferring
the pVLT31 vector harboring flavin-oxidoreductase gene (dszD) into
the recombinant P. aeruginosa ATCC 9027 which contained dszABC
gene in its chromosome stably [52]. This recombinant bacteria
showed the enhanced activity of biodesulfurization when all four
genes co expressed. The biodesulfurization efficiency has also been
significantly improved using the approach of directed evolution.
Arensdorf et al. (2002) presented an example of directed evolution
of biodesulfurization using chemostat approach [53]. Rhodococcus
erythropolis IGTS8 has not ability to utilize octyl sulphide and
5-methylbenzothiophene (5-MBT) as sole sulphur sources. Arensdorf
et al. (2002) selected gain-of-function mutants of strain IGTS8 using
sulphur limited chemostat [53]. These mutants were grown with octyl
sulphide and 5-MBT as sole sulphur sources. The ability of mutant
to grow on 5-MBT as sole sulphur source was due to a transversion
(guanine to thymine) in dszC codon 261 [53].

Monooxygenases and Denitrification of Nitro Aromatic

Compounds
Nitroaromatic compounds are serious environment pollutants
that are used in the synthesis of pesticides, drugs, dyes and explosives
[54]. Nitro phenols are the most common examples of nitroaromatic
compounds. The toxicity of these compounds is mainly due to the
presence of nitro group in the aromatic ring. The aerobic degradation

of these compounds generally initiate with removal of the nitro group

by oxygenase activity [55]. Examples of monooxygenases involved
in denitrification of nitroaromatic compounds are 4-nitrophenol
4-monooxygenase,
4-nitrophenol
2-monooxygenase
and
2-nitrophenol 2-monooxygenase. 4-Nitrophenol 4-monooxygenase
(EC 1.14.13.-) has been purified and characterized from Pseudomonas
sp. strain WBC-3 [56]. This enzyme is a single component and flavin
adenine dinucelotide monooxygenase that catalyzes conversion of
p-nitrophenol to p-benzoquinone in the presence of NADPH (Figure
4a). This enzyme also catalyzes NADPH dependent conversion of
nitrocatechol to 1, 2, 4-benzenetriol [57]. Another denitrifying
4-nitrophenol 2-monooxygenase (EC 1.14.13.29) has been purified
and characterized from Rhodococcus sp. strain PN1 [58]. This enzyme
is a two-component flavin adenine dinucelotide monooxygenase
and converts p-nitrophenol to 4-nitrocatechol in the presence
of NADH (Figure 4b) whereas NADPH dependent 2-nitrophenol2-monooxygenase (EC 1.14.13.31) catalyzes the conversion of
2-nitrophenol to catechol (Figure 4c) [55].

Monooxygenases and hydroxylation of aromatic compounds

Several monooxygenases have been identified and characterized
that involved in the biodegradation of various aromatic compounds
and have been listed in the (Table 1). A few monooxygenases acting on
aromatic compounds have been used as biocatalysis for synthesis of
the pharmaceuticals compounds. Examples of these monooxygenases
are styrene monooxygenase, hydroxybiphenyl 3-monooxygenase
and phenylacetone monooxygenase. Styrene monooxygenase (EC
1.14.13.-) from Pseudomonas sp. VLB 120 has ability to convert
styrene to s-styrene oxide with an enantiomeric excess of greater than

Enzyme

E.C number

Substrate(s)

References

4-Methyl 5-nitrocatechol monooxygenase

1.13.12.-

4-Methyl-5-nitrocatechol

[58]

Toluene 3-monooxygenase

1.14.-.-

3-Hydroxytoluene, Toluene

[59]

Toluene 2-monooxygenase

1.14.13.-

2-Hydroxytoluene, Toluene, 1,1,2-Trichloroethylene [60]

4-Hydroxyacetophenone monooxygenase

1.14.13.-

4-Hydroxyacetophenone

[61]

Toluene 4-monooxygenase

1.14.13.-

Toluene, N-Nitrosodimethylamine

[62]

4-Hydroxybenzoate 3-monooxygenase

1.14.13.2

4-Hydroxybenzoate

[63]

2-Mercaptobenzothiazole monooxygenase

http://umbbd.msi.umn.edu/servlets/
pageservlet?ptype=r&reacID=r1178 1.14.-.-

2-Mercaptobenzothiazole

[64]

6-Hydroxy-2-mercaptobenzothiazole
monooxygenase

1.14.-.-

6-Hydroxy-2-mercaptobenzothiazole

[64]

Benzothiazole monooxygenase

1.14.-.-

Benzothiazole

[65]

Phenylboronic acid monooxygenase

1.14.-.-

Phenylboronic acid

[66]

2,6-Dihydroxybenzothiazole monooxygenase

1.14.-.-

2,6-Dihydroxybenzothiazole

[67]

2-Hydroxybenzothiazole monooxygenase

1.14.-.-

2-Hydroxybenzothiazole

[65]

1-Indanone monooxygenase

1.14.-.-

1-Indanone

[68]

2-Indanone monooxygenase

1.14.-.-

2-Indanone

[68]

6-Hydroxy-3-methyl-2-oxo-1,2-dihydroquinoline
6-monooxygenase

1.14.-.-

6-Hydroxy-3-methyl-2-oxo-1,2-dihydroquinoline

[69]

Toluene-4-sulfonate monooxygenase

1.14.-.-

Toluene 4-sulfonate

[70]

Dibenzothiophene 5,5-dioxide monooxygenase

1.14.13.-

Dibenzothiophene-5,5-dioxide

[71]

3-Methyl 2-oxo 1,2-dihydroquinoline

6-monooxygenase

1.14.-.-

3-Methyl-2-oxo-1,2-dihydroquinoline

[69]

BCDS monooxygenase

1.14.-.-

Branched-chain dodecylbenzene sulfonate

[72]

Toluene sulfonate methyl monooxygenase

1.14.13.-

Toluene-4-sulfonate

[73]

Orcinol 2-monooxygenase

1.14.13.6

Orcinol

[74]

3-Hydroxybenzoate 4-hydroxylase

1.14.13.23

3-Hydroxybenzoate

[75]

3-Hydroxybenzoate 6-monooxygenase

1.14.13.24

3-Hydroxybenzoate

[76]

4-Methoxybenzoate monooxygenase

1.14.99.15

4-methoxybenzoate

[77]

Benzoate 4-monooxygenase

1.14.13.12

Benzoate

[78]

N-isopropylacetaniline monooxygenase

1.14.15.-

N-Isopropylacetanilide

[79]

Volume 1 Issue 31000112

lishing
Pub
Gr
CS

p
ou

J Bioremed Biodegrad
ISSN:2155-6199 JBRBD, an open access journal

OM
I

Table 1: Monooxygen6+ases involved in the biodegradation of aromatic compounds.

Citation: Arora PK, Srivastava A, Singh VP (2010) Application of Monooxygenases in Dehalogenation, Desulphurization, Denitrification and
Hydroxylation of Aromatic Compounds. J Bioremed Biodegrad 1:112. doi:10.4172/2155-6199.1000112

Page 5 of 8

Phenylacetone monooxygenase (PAMO, EC 1.14.13.92) is a FADcontaining Baeyer-Villiger monooxygenase (BVMO). This enzyme was
discovered from Thermobifida fusca that is a moderately thermophilic
soil bacterium and grows at 55C. PAMO oxidizes phenylacetone,
aromatic and aliphatic ketones, aromatic sulphides and sulphoxides
and organoborn compounds [87]. This enzyme has thermostability
and tolerance towards organic solvents [88], therefore, this enzyme
is suitable for various industrial applications.

Current ways to imporve the efficiency of monooxygenases

Metagenomic approach: Microbial diversity is a rich source
of novel enzymes including monooxygenases. To date, only 1%
microorganisms could be cultured in the laboratory conditions and
monooxygenases associated with these culturable microbes have
been identified and characterized. Most potential monooxygenases
remain untapped because vast majority of bacteria are uncluturable.
In such a case, metagenomics may be powerful tool to explore
novel monooxygenases from entire community of microbes. The
metagenomic approach involves (i) the isolation and purification of
DNA from an environmental sample, (ii) cloning of DNA fragments into
suitable vectors, (iii) the transformation of host cells with construct
and (iv) functional and sequence based screening of constructed
clones. Numerous novel biocatalysts have been isolated from various
metagenomic libraries. A novel styrene monooxygenase (SmoA) was
discovered by the screening of a metagenomic library from loam
soil [89]. SmoA is highly enantioselective, since it is able to convert
aromatic alkenes into the (S)-epoxides with an excellent enantiomeric
excess. Recently, two flavin monooxygenases catalyzing the oxidation
of indole to a mixture of indigo and indirubin pigments have been
identified from an effluent treatment plant sludge metagenomic
library. A new self-sufficient CYP, SYK181was identified during
sequence-based screening of a metagenomic library [91]. SYK181
showed significant hydroxylase activity towards naphthalene and
phenanthrene as well as towards fatty acids [91]. Therefore, it is very
useful for the biodegradation of organic chemicals.
Directed evolution: In the recent years, directed evolution is
playing a major role for engineering of the proteins for bio catalytic
applications. Directed evolution is based on the Darwinian principle
of evolution and involves (i) random mutagenesis to generate the
diversity and (ii) the subsequent selection of the improved variants
by screening. The efficiency of several monooxygenases has been
significantly improved by directed evolution. For example, CYP102
from Bacillus megaterium BM3 has been turned by directed
evolution into an enzyme that hydrolyze aromatic compounds
like 2,6-dichlorophenol and 2-benzyloxyphenol which cannot be
hydrolyzed by wild type enzyme [92]. Furthermore, a triple mutant

Bioinformatic approach: Database providing information about

the biochemistry and genetics of monooxygenases are valuable tools
of the computational biology. Currently, two databases namely UMBBD (University of Minnesota Biocatalysis/Biodegradation Database)
[100,101] and OXDBase (A Database of Biodegradative Oxygenases)
[1] are providing the information about all monooxygenases involved
in biodegradation. The characteristics feature of these databases is
to provide information of the genes and three dimensional structures
of the monooxygenases which can help in site directed mutagenesis
of the monooxygenases to improve their catalytic properties [1].
The entries of the monooxygenases in OxDBase as well as UM-BBD
are linked to various existing databases which provide to users the
detailed information of monooxygenases [1]. Since monooxygenases
catalyzed biotransformations of the toxic xenobiotic compounds help
in reducing the toxicity of the xenobiotics, therefore, the detailed
information of these monooxygenases increase our understanding of
biodegradation process [1].
Homology modelling is a tool of bioinformatics that use to
generate reasonable models of protein structures. The determination
of 3D structure of monooxygenase by homology modelling is a helpful
to design the experiment of site directed mutagenesis in active site
of that protein to improve the efficiency of the monooxygenase.
The three dimensional structure of tolune 4monooxygenase
determined by homology modelling has helped in altering the
activity of this enzyme by active site engineering for the synthesis of
3-methoxycatechol, methoxyhydroquinone and methylhydroquinone
[102].

Conclusion
Monooxygenases are multifunctional enzymes and involved in
biodesulfurization, dehalogenation, denitrification and hydroxylation
of various aromatic compounds. The applications of monooxygenases
have been improved using the recent molecular and bioinformatic
approaches.
References
1. Arora PK, Kumar M, Chauhan A, Raghava GP, Jain RK (2009) OxDBase: a
database of oxygenases involved in biodegradation. BMC Res Notes 2: 67.
2. van Berkel WJH, Kamerbeek NM, Fraaije MW(2006) Flavoprotein
monooxygenases, a diverse class of oxidative biocatalysts. J Biotechnol 124:
670-689.
3. Urlacher VB, Lutz-Wahl S, Schmid RD (2004) Microbial P450 enzymes in
biotechnology. Appl Microbiol Biotechnol 64: 317-325.
4. Pazmino DET, Winkler M, Glieder A, Fraaije MW (2010) Monooxygenases
as biocatalysts: Classification, mechanistic aspects and biotechnological
applications. J Biotechnol 146: 9-24.

Volume 1 Issue 31000112

lishing
Pub
Gr
CS

p
ou

J Bioremed Biodegrad
ISSN:2155-6199 JBRBD, an open access journal

of CYP102A1 has converted -ionone into flavorant (R)-4-hydroxy-ionone in a regioselective manner and exhibited 80-fold higher activity
compared to the wild type enzyme [93] . Another example of directed
evolution is cyclohexanone monooxygenase (CHMO; EC 1.14.13.22)
from Acinetobacter sp. NCIMB 9871. This enzyme act as a biocatalyst
in the BaeyerVilliger (BV) reaction of 4-hydroxycyclohexanone and
other 4-substituted cyclohexanone derivatives [94]. It has been
used for commercial production of bicyclic lactone, sulfoxides,
thiosulfinates and cyclic sulphates [95-97]. The range of the
substrates and enantioselectivity of this enzyme have been increased
by directed evolution [98,99]. Reetz et al. (2004) demonstrated that
some CHMO mutants showed improved enantioselectivity, preferring
the (R)-enantiomer (49-54% ee vs 9% ee for the wild type enzyme)
in the oxidation of prochiral 4-hydroxycyclohexanone whereas other
variants favored (S)-enantiomer (79% ee).

OM
I

99% [81]. Styrene monooxygenase is a two component enzyme: StyA

(oxygenase) and StyB (reductase). A recombinant E. Coli expressing
StyA and StyB has been used as whole cell biocatalyst for a broad
substrate spectrum that gives access to various chiral acryloxides
[82]. Another commercially important enzyme, 2-hydroxybiphenyl
3-monooxygenase (HbpA, EC 1.14.13.44) from Pseudomonas azelaica
HBP1 catalyzes ortho-hydroxylation of 2-hydroxybiphenyl to 2,
3-dihydroxybiphenyl and also catalyzes the conversion of different
2-substituted phenols to 3-substituted catechols that are of interests
of pharmaceuticals industry [83,84] . The substrate reactivity of
2-hydroxybiphenyl 3-monooxygenase (HbpA) have been improved
by directed evolution [84]. A variant of recombinant E. coli JM101
expressing HbpA was used for the production of 3-tert-butylcatechol,
a compound of pharmaceutical interest [86].

Citation: Arora PK, Srivastava A, Singh VP (2010) Application of Monooxygenases in Dehalogenation, Desulphurization, Denitrification and
Hydroxylation of Aromatic Compounds. J Bioremed Biodegrad 1:112. doi:10.4172/2155-6199.1000112

Page 6 of 8
5. Shen B, Hutchinson CR (1993) Tetracenomycin F1 monooxygenase:
Oxidation of a naphthacenone to a naphthacenequinone in the biosynthesis of
tetracenomycin C in Streptomyces glaucescens. Biochemistry 32: 6656-6663.

27. Martin-Le Garrec G, Artaud I, Capeillere-Blandin C (2001) Purification and

catalytic properties of the chlorophenol 4-monooxygenase from Burkholderia
cepacia strain AC1100. Biochim Biophys Acta 1547: 288-301.

6. Fetzner S (2002) Oxygenases without requirement for cofactors or metal ions.

Appl Microbiol Biotechnol 60:243-257.

28. Webb BN, Ballinger JW, Kim E, Belchik SM, Lam KS, et al. (2010)
Characterization of Chlorophenol 4-Monooxygenase (TftD) and NADH:FAD
Oxidoreductase (TftC) of Burkholderia cepacia AC1100. J Bio Chem 285:20142027.

8. Lemieux MJ, Ference C, Cherney MM, Wang M, Garen C, et al. (2005)

The crystal structure of Rv0793, a hypothetical monooxygenase from M.
tuberculosis. J Struct Funct Genomics 6: 245-257.
9. Bertini I, Briganti F, Mangani S, Nolting HF, Scozzafava A (1995) Biophysical
investigation of bacterial aromatic extradiol dioxygenases involved in
biodegradation processes. Coordin Chem Rev 144: 321-345.
10. Boyd DR, Sharma ND, Allen CC (2001) Aromatic dioxygenases: molecular
biocatalysis and applications. Curr Opin Biotechnol 12: 564-573.
11. Gibson DT, Parales RE (2000) Aromatic hydrocarbon dioxygenases in
environmental biotechnology. Curr Opin Biotechnol 11: 236-243.
12. Lipscomb JD (2008) Mechanism of extradiol aromatic ring cleaving
dioxygenases. Curr Opin Struct Biol 18: 644-649.
13. Crawford RL, Jung CM, Strap JL (2007) The recent evolution of
pentachlorophenol (PCP)-4-monooxygenase (PcpB) and associated pathways
for bacterial degradation of PCP. Biodegradation 18: 525-539.
14. Copley SD (1997) Diverse mechanistic approaches to difficult chemical
transformations: Microbial dehalogenation of chlorinated aromatic compounds.
Chem Biol 4: 169-174.
15. Lange CC, Schneider BJ, Orser CS (1996) Verification of the role of PCP
4-monooxygenase in chlorine elimination from pentachlorophenol by
Flavobacterium sp. strain ATCC 39723. Biochem Biophys Res Commun 219:
146-149.
16. Xun L, Topp E, Orser CS (1992) Diverse substrate range of a Flavobacterium
pentachlorophenol hydroxylase and reaction stoichiometries. J Bacteriol 174:
2898-2902.
17. Orser CS, Lange CC, Xun L, Zahrt TC, Schneider BJ (1993) Cloning,
sequence analysis, and expression of the Flavobacterium pentachlorophenol
4-monooxygenase gene in Escherichia coli. J Bacteriol 175: 411-416.
18. Saboo VM, Gealt MA (1998) Gene sequences of the pcpB gene of
pentachlorophenol-degrading Sphingomonas chlorophenolica found in
nondegrading bacteria. Can J Microbiol 44: 667-675.
19. Karlson U, Rojo F, vanElsas JD, Moore E (1996) Genetic and serological
evidence for the recognition of four pentachlorophenol degrading bacterial
strains as a species of the genus Sphingomonas. System Appl Microbiol 18:
539-548.
20. Ederer MM, Crawford RL, Herwig RP, Orser CS (1997) PCP degradation is
mediated by closely related strains of the genus Sphingomonas. Mol Ecol 6:
39-49.
21. Cassidy MB, Lee H, Trevors JT, Zablotowicz RB (1999) Chlorophenol and
nitrophenol metabolism by Sphingomonas sp UG30. J Ind Microbiol Biotechnol
23: 232-241.
22. Leung KT, Campbell S, Gan Y, White DC, Lee H, Trevors JT (1999) The role
of the Sphingomonas species UG30 pentachlorophenol-4-monooxygenase in
p-nitrophenol degradation. FEMS Microbiol Lett 173: 247-253.
23. Leung KT, Cassidy MB, Shaw KW, Lee H, Trevors JT, et al. (1997)
Pentachlorophenol biodegradation by Pseudomonas spp. UG25 and UG30.
World J Microbiol Biotechnol 13: 305-313.
24. Tiirola MA, Mannisto MK, Puhakka JA, Kulomaa MS (2002) Isolation
and characterization of Novosphingobium sp. strain MT1, a dominant
polychlorophenol-degrading strain in a groundwater bioremediation system.
Appl Environ Microbiol 68: 173-180.
25. Wang H, Marjomaki V, Ovod V, Kulomaa MS (2002) Subcellular localization
of pentachlorophenol 4-monooxygenase in Sphingobium chlorophenolicum
ATCC 39723. Biochem Biophys Res Commun 299: 703-709.
26. Nakamura T, Motoyama T, Hirono S, Yamaguchi I (2004) Identification,
characterization,
and
site-directed
mutagenesis
of
recombinant
pentachlorophenol 4-monooxygenase. Biochim Biophys Acta 1700: 151-159.

30. Xun L (1996) Purification and characterization of chlorophenol 4-monooxygenase

from Burkholderia cepacia AC1100. J Bacteriol 178: 2645-2649.
31. Wieser M, Wagner B, Eberspacher J, Lingens F (1997) Purification and
characterization of 2,4,6-trichlorophenol-4-monooxygenase, a dehalogenating
enzyme from Azotobacter sp strain GP1. J Bacteriol 179:202-208.
32. Xun L, Webster CM (2004) A monooxygenase catalyzes sequential
dechlorinations of 2,4,6-trichlorophenol by oxidative and hydrolytic reactions.
J Biol Chem 279:6696-6700.
33. Kilbane JJ, Jackowski K (1992) Biodesulfurization of water soluble coal derived
material by Rhodococcus rhodochrous IGTS8. Biotechnol Bioeng 40: 11071114.
34. Denome SA, Oldfield C, Nash LJ, Young KD (1994) Characterization of the
desulfurization genes from Rhodococcus sp. strain IGTS8. J Bacteriol 176:
6707-6716.
35. Li MZ, Squires CH, Monticello DJ, Childs JD (1996) Genetic analysis of the
dsz promoter and associated regulatory regions of Rhodococcus erythropolis
IGTS8. J Bacteriol 178: 6409-6418.
36. DenisLarose C, Labbe D, Bergeron H, Jones AM, Greer CW, et al, (1997)
Conservation of plasmid-encoded dibenzothiophene desulfurization genes in
several Rhodococci. Appl Environ Microbiol 63: 2915-2919.
37. Omori T, Monna L, Saiki Y, Kodama T (1992) Desulfurization of dibenzothiophene
by Corynebacterium sp. strain-Sy1. Appl Environ Microbiol 58: 911-915.
38. Izumi Y, Ohshiro T, Ogino H, Hine Y, Shimao M (1994) Selective desulfurization
of dibenzothiophene by Rhodococcus erythropolis D-1. Appl Environ Microbiol
60: 223-226.
39. Omori T, Saiki Y, Kasuga K, Kodama T (1995) Desulfurization of alkyl
and aromatic sulfides and sulfonates by dibenzothiophene-desulfurizing
Rhodococcus sp. strain Sy1. Biosci Biotechnol Biochem 59: 1195-1198.
40. Wang P, Krawiec S (1994) Desulfurization of dibenzothiophene to
2-hydroxybiphenyl by some newly isolated bacterial strains. Arch Microbiol
161: 266-271.
41. Shavandi M, Sadeghizadeh M, Khajeh K, Mohebali G, Zomorodipour A
(2010) Genomic structure and promoter analysis of the dsz operon for
dibenzothiophene biodesulfurization from Gordonia alkanivorans RIPI90A.
Appl Microbiol Biotechnol 87: 1455-1461.
42. Kilbane JJ, Robbins J (2007) Characterization of the dszABC genes of
Gordonia amicalis F.5.25.8 and identification of conserved protein and DNA
sequences. Appl Microbiol Biotechnol 75: 843-851.
43. Xiong XC, Li WL, Li X, Xing JM, Liu HZ (2005) Comparison of the desulfurization
activity among several bacteria and analysis of the conservation of their
desulfurization genes. Wei Sheng Wu Xue Bao 45: 733-737.
44. Kirimura K, Harada K, Iwasawa H, Tanaka T, Iwasaki Y, et al. (2004) Identification
and functional analysis of the genes encoding dibenzothiophene-desulfurizing
enzymes from thermophilic bacteria. Appl Microbiol Biotechnol 65: 703-713.
45. Srinivasaraghavan K, Sarma PM, Lal B (2006) Comparative analysis of
phenotypic and genotypic characteristics of two desulfurizing bacterial strains,
Mycobacterium phlei SM120-1 and Mycobacterium phlei GTIS10. Lett Appl
Microbiol 42: 483-489.
46. Furuya T, Kirimura K, Kino K, Usami S (2001) Thermophilic biodesulfurization
of dibenzothiophene and its derivatives by Mycobacterium phlei WU-F1. FEMS
Microbiol Lett 204: 129-133.
47. Konishi J, Onaka T, Ishii Y, Suzuki M (2000) Demonstration of the carbon-sulfur
bond targeted desulfurization of benzothiophene by thermophilic Paenibacillus
sp. strain A11-2 capable of desulfurizing dibenzothiophene. FEMS Microbiol
Lett 187: 151-154.

Volume 1 Issue 31000112

lishing
Pub
Gr
CS

p
ou

J Bioremed Biodegrad
ISSN:2155-6199 JBRBD, an open access journal

29. Gisi MR, Xun LY (2003) Characterization of chlorophenol 4-monooxygenase

(TftD) and NADH : flavin adenine dinucleotide oxidoreductase (TftC) of
Burkholderia cepacia AC1100. J Bacteriol 185: 2786-2792.

OM
I

7. Adams MA, Jia Z (2005) Structural and biochemical evidence for an enzymatic
quinone redox cycle in Escherichia coli: identification of a novel quinol
monooxygenase. J Biol Chem 280: 8358-8363.

Citation: Arora PK, Srivastava A, Singh VP (2010) Application of Monooxygenases in Dehalogenation, Desulphurization, Denitrification and
Hydroxylation of Aromatic Compounds. J Bioremed Biodegrad 1:112. doi:10.4172/2155-6199.1000112

Page 7 of 8
48. Raheb J, Hajipour MJ, Saadati M, Rasekh B, Memari B (2009) The enhancement
of biodesulfurization activity in a novel indigenous engineered Pseudomonas
putida. Iran Biomed J 13: 207-213.

69. Casellas M, Grifoll M, Bayona JM, Solanas AM (1997) New metabolites in the
degradation of fluorene by Arthrobacter sp. strain F101. Appl Environ Microbiol
63:819-826.

49. Shavandi M, Sadeghizadeh M, Zomorodipour A, Khajeh K (2009)

Biodesulfurization of dibenzothiophene by recombinant Gordonia alkanivorans
RIPI90A. Bioresour Technol 100: 475-479.

70. Schach S, Schwarz G, Fetzner S, Lingens F (1993) Microbial metabolism of

Quinoline and related compounds. XVII. Degradation of 3-methylquinoline by
Comamonas testosteroni 63. Biol Chem Hoppe-Seyler 374: 175-181.

50. Gallardo ME, Ferrandez A, DeLorenzo V, Garcia JL, Diaz E (1997) Designing
recombinant Pseudomonas strains to enhance biodesulfurization. J Bacteriol
179:7156-7160.

71. Beil S, Kertesz MA, Leisinger T, Cook AM (1996) The assimilation of sulfur from
multiple sources and its correlation with expression of the sulfate-starvationinduced stimulon in Pseudomonas putida S-313. Microbiology 142: 1989-1995.

51. Calzada J, Zamarro MT, Alcon A, Santos VE, Diaz E, et al. (2009) Analysis of
dibenzothiophene desulfurization in a recombinant Pseudomonas putida strain.
Appl Environ Microbiol 75: 875-877.

72. Oldfield C, Pogrebinsky O, Simmonds J, Olson ES, Kulpa CF (1997)

Elucidation of the metabolic pathway for dibenzothiophene desulphurization
by Rhodococcus sp. strain IGTS8 (ATCC 53968). Microbiol 143: 2961-2973.

52. Raheb J, Hajipour MJ, Memari B (2010) Increasing of biodesulfurization activity

of newly recombinant Pseudomonas aeruginosa ATCC 9027 by cloning the
flavin reductase gene. Inter J Biotechnol Biochem 6: 219-229.

73. Campos-Garcia J, Esteve A, Vazquez-Duhalt R, Ramos JL, Soberon-Chavez G

(1999) The branched chain dodecylbenzene sulfonate degradation pathway of
Pseudomonas aeruginosa W51D involves a novel route for degradation of the
surfactant lateral alkyl chain. Appl Environ Microbiol 65: 3730-3734.

53. Arensdorf JJ, Loomis AK, DiGrazia PM, Monticello DJ, Pienkos PT (2002)
Chemostat approach for the directed evolution of biodesulfurization gain-offunction mutants. Appl Environ Microbiol 68: 691-698.
54. Ju KS, Parales RE (2010) Nitroaromatic compounds, from synthesis to
biodegradation. Microbiol Mol Biol Rev 74: 250-272.
55. Zeyer J, Kocher HP (1988) Purification and characterization of a bacterial
nitrophenol oxygenase which converts ortho-nitrophenol to catechol and nitrite.
J Bacteriol 170: 1789-1794.
56. Zhang JJ, Liu H, Xiao Y, Zhang XE, Zhou NY (2009) Identification and
characterization of catabolic para-nitrophenol 4-monooxygenase and parabenzoquinone reductase from Pseudomonas sp. strain WBC-3. J Bacteriol
191: 2703-2710.

74. Locher HH, Leisinger T, Cook AM (1991) 4-Toluene sulfonate methyl

monooxygenase from Comamonas testosteroni T-2: Purification and some
properties of the oxygenase component. J Bacteriol 173: 3741-3748.
75. Chapman PJ, Ribbons DW (1976) Metabolism of resorcinylic compounds by
bacteria: orcinol pathway in Pseudomonas putida. J Bacteriol 125: 975-984.
76. Nakazawa T, Hayashi E (1978) Phthalate and 4-hydroxyphthalate
metabolism in Pseudomonas testosteroni: purification and properties of
4,5-dihydroxyphthalate decarboxylase. Appl Environ Microbiol 36: 264-269.
77. Poh CL, Bayly RC (1980) Evidence for isofunctional enzymes used in m-cresol
and 2,5-xylenol degradation via the gentisate pathway in Pseudomonas
alcaligenes. J Bacteriol 143: 59-69.

57. Wei M, Zhang JJ, Liu H, Zhou NY (2010) para-Nitrophenol 4-monooxygenase

and hydroxyquinol 1,2-dioxygenase catalyze sequential transformation of
4-nitrocatechol in Pseudomonas sp. strain WBC-3. Biodegradation 21: 915921

78. Bernhardt FH, Bill E, Trautwein AX, Twilfer H (1988) 4-Methoxybenzoate

monooxygenase from Pseudomonas putida: isolation, biochemical properties,
substrate specificity, and reaction mechanisms of the enzyme components.
Methods Enzymol 161: 281-294.

58. Takeo M, Murakami M, Niihara S, Yamamoto K, Nishimura M, et al (2008)

Mechanism of 4-Nitrophenol oxidation in Rhodococcus sp. strain PN1:
characterization of the two-component 4-nitrophenol hydroxylase and
regulation of Its expression. J Bacteriol 190: 7367-7374.

79. Sahasrabudhe SR, Modi VV (1995) Hydroxylation of benzoate and its

chlorinated derivatives in Aspergillus niger. Biochem Int 10: 525-529.

59. Leungsakul T, Johnson GR, Wood TK (2006) Protein engineering of the

4-methyl-5-nitrocatechol monooxygenase from Burkholderia sp. strain DNT for
enhanced degradation of nitroaromatics. Appl Environ Microbiol 72: 3933-3939.

80. Martin M, Mengs G, Plaza E, Garbi C, Sanchez M, et al. (2000) Propachlor

removal by Pseudomonas strain GCH1 in an immobilized cell system. Appl
Environ Microbiol 66: 1190-1194.

60. Olsen RH, Kukor JJ, Kaphammer B (1994) A novel Toluene 3-monooxygenase
pathway cloned from Pseudomonas pickettii PKO1. J Bacteriol176: 3749-3756.

81. Panke S, Witholt B, Schmid A, Wubbolts MG (1998) Towards a biocatalyst

for (S)-styrene oxide production: characterization of the styrene degradation
pathway of Pseudomonas sp. strain VLB120. Appl Environ Microbiol 64:20322043.

61. Newman LM, Wackett LP (1995) Purification and characterization of toluene

2-monooxygenase from Burkholderia cepacia G4. Biochemistry 34:1406614076.

82. Schmid A, Feiten HJ, Hollmann F, Witholt B (2001) Integrated biocatalytic

synthesis on gram scale: the highly enantioselective preparation of chiral
oxiranes with styrene monooxygenase. Adv Synth Catal 343: 732737.

62. Rehdorf J, Zimmer CL, Bornscheuer UT (2009) Cloning, expression,

characterization, and biocatalytic investigation of the 4-hydroxyacetophenone
monooxygenase from Pseudomonas putida JD1. Appl Environ Microbiol 75:
3106-3114.

83. Held M, Schmid A, Kohler HPE, Suske W, Witholt B, et al. (1999) An integrated
process for the production of toxic catechols from toxic phenols based on a
designer biocatalyst. Biotechnol Bioeng 62: 641-648.

63. Oppenheim SF, Studts JM, Fox BG, Dordick JS (2001) Aromatic hydroxylation
catalyzed by toluene 4-monooxygenase in organic solvent/aqueous buffer
mixtures. Appl Biochem Biotechnol 90: 187-197.

84. Held M, Suske W, Schmid A, Engesser KH, Kohler HPE, et al. (1998) Preparative
scale production of 3-substituted catechols using a novel monooxygenase from
Pseudomonas azelaica HBP 1. J Mol Catal B-Enzym 5: 87-93.
85. Meyer A, Schmid A, Held M, Westphal AH, Rothlisberger M, et al. (2002)
Changing the substrate reactivity of 2-hydroxybiphenyl 3-monooxygenase from
Pseudomonas azelaica HBP1 by directed evolution. J Biol Chem 277: 55755582.

65. Haroune N, Combourieu B, Besse P, Sancelme M, Kloepfer A, et al. (2004)

Metabolism of 2-mercaptobenzothiazole by Rhodococcus rhodochrous. Appl
Environ Microbiol 70: 6315-6319.

86. Meyer A, Held M, Schmid A, Kohler HP, Witholt B (2003) Synthesis of 3-tertbutylcatechol by an engineered monooxygenase. Biotechnol Bioeng 81:518524.

66. Besse P, Combourieu B, Boyse G, Sancelme M, De Wever H, et al. (2001)

Long-range (1)H-(15)N heteronuclear shift correlation at natural abundance: a
tool to study benzothiazole biodegradation by two Rhodococcus strains. Appl
Environ Microbiol 67: 1412-1417.

87. Pazmino DET, de Gonzalo G, Ottolina G, Carrea G, Janssen DB, et al.

(2005) Exploring the biocatalytic potential of the novel thermostable BaeyerVilliger monooxygenase: Phenylacetone monooxygenase. J Biotechnol 118:
S129-S129.

67. Negrete-Raymond AC, Weder B, Wackett LP (2003) Catabolism of arylboronic

acids by Arthrobacter nicotinovorans strain PBA. Appl Environ Microbiol 69:
4263-4267.

88. de Gonzalo G, Pazmino DET, Ottolina G, Fraaije MW, Carrea G (2005)

Oxidations catalyzed by phenylacetone monooxygenase from Thermobifida
fusca. Tetrahedron Asymmetr 16: 3077-3083.

68. Haroune N, Combourieu B, Besse P, Sancelme M, Reemtsma T, et al. (2002)

Benzothiazole degradation by Rhodococcus pyridinovorans strain PA: evidence
of a catechol 1,2-dioxygenase activity. Appl Environ Microbiol 68: 6114-6120.

89. van Hellemond EW, Janssen DB, Fraaije MW (2007) Discovery of a novel
styrene monooxygenase originating from the metagenome. Appl Environ
Microbiol 73: 5832-5839.

Volume 1 Issue 31000112

lishing
Pub
Gr
CS

p
ou

J Bioremed Biodegrad
ISSN:2155-6199 JBRBD, an open access journal

OM
I

64. Salituro FG, Demeter DA, Weintraub HJ, Lippert BJ, Resvick RJ, et al. (1994)
Multisubstrate inhibition of 4-hydroxybenzoate 3-monooxygenase. J Med
Chem 37: 4076-4078.

Citation: Arora PK, Srivastava A, Singh VP (2010) Application of Monooxygenases in Dehalogenation, Desulphurization, Denitrification and
Hydroxylation of Aromatic Compounds. J Bioremed Biodegrad 1:112. doi:10.4172/2155-6199.1000112

Page 8 of 8
90. Singh A, Singh Chauhan N, Thulasiram HV, Taneja V, Sharma R (2010)
Identification of two flavin monooxygenases from an effluent treatment plant
sludge metagenomic library. Bioresour Technol 101: 8481-8484.

97. Colonna S, Gaggero N, Carrea G, Pasta P (1998) Oxidation of organic

cyclic sulfites to sulfates: a new reaction catalyzed by cyclohexanone
monooxygenase. Chem Commun 3: 415-416.

91. Kim BS, Kim SY, Park J, Park W, Hwang KY, et al. (2007) Sequence-based
screening for self-sufficient P450 monooxygenase from a metagenome library.
J Appl Microbiol 102: 1392-1400.

98. Mihovilovic MD, Rudroff F, Winninger A, Schneider T, Schulz F, et al. (2006)

Microbial Baeyer-Villiger oxidation: Stereopreference and substrate acceptance
of cyclohexanone monooxygenase mutants prepared by directed evolution.
Organic Letters 8: 1221-1224.

93. Urlacher VB, Makhsumkhanov A, Schmid RD (2006) Biotransformation of betaionone by engineered cytochrome P450BM-3. Appl Microbiol Biotechnol 70:
53-59.
94. Mihovilovic MD, Muller B, Stanetty P (2002) Monooxygenase-mediated BaeyerVilliger oxidations. Eur J Org Chem 22: 3711-3730.
95. Ottolina G, de Gonzalo G, Carrea G, Danieli B (2005) Enzymatic Baeyer-Villiger
oxidation of bicyclic diketones. Adv Synth Catal 347: 1035-1040.
96. Colonna S, Del Sordo S, Gaggero N, Carrea G, Pasta P (2002) Enzymemediated catalytic asymmetric oxidations. Heteroatom Chem 13: 467-473.

100. Gao JF, Ellis LBM, Wackett LP (2010) The University of Minnesota Biocatalysis/
Biodegradation Database: improving public access. Nucleic Acids Res 38:
D488-D491.
101. Arora P, Shi W (2010) Tools of bioinformatics in biodegradation. Revi Environ
Sci Biotechnol 9: 211-213.
102. Tao Y, Fishman A, Bentley WE, Wood TK (2004) Altering toluene
4-monooxygenase by active-site engineering for the synthesis of
3-methoxycatechol, methoxyhydroquinone and methylhydroquinone. J
Bacteriol 186: 4705-4713.

Volume 1 Issue 31000112

lishing
Pub
Gr
CS

p
ou

J Bioremed Biodegrad
ISSN:2155-6199 JBRBD, an open access journal

99. Reetz MT, Brunner B, Schneider T, Schulz F, Clouthier CM, et al. (2004)
Directed evolution as a method to create enantioselective cyclohexanone
monooxygenases for catalysis in Baeyer-Villiger reactions. Angew Chem Int
Engl 43: 4075-4078.

OM
I

92. Sulistyaningdyah WT, Ogawa J, Li QS, Maeda C, Yano Y, et al. (2005)

Hydroxylation activity of P450 BM-3 mutant F87V towards aromatic compounds
and its application to the synthesis of hydroquinone derivatives from phenolic
compounds. Appl Microbiol Biotechnol 67: 556-562.