Microbial epoxide hydrolase

1. Microbial Epoxide Hydrolase
By
Saima Riaz (Ph.D Scholar)

2. Epoxides
• Organic, three membered heterocyclic compound of oxygen and
carbon
• Unstable in water
• Generate in various metabolic process like during the metabolism
of xenobiotic compounds ( the foreign compound to the body such
as food, drugs and pollutant as well as produced inside body as a
metabolite of various process like steroids, bile acid and certain
fatty acids)
• They are also called epoxy, epoxide, oxirane, and ethoxyline. Simple
epoxides are often referred to as oxides

3. Role of Epoxide
• Used as signal molecule
• Defensive agent
• Blood vessel relaxation
• Inflammatory process
• DNA modification
• Mutation
• carcinogens

4. Epoxide Hydrolases
• An enzyme controlled the over production of Epoxide
• Identified in many organism like Mammals,
plants, insects and microbes
• Distinct type of EHs
• Soluble epoxide hydrolase (sEH)
• microsomal epoxide hydrolase (mEH)
• limonene-1,2-epoxide hydrolase (LEH)
• Many others

5. Microbial Epoxide Hydrolase
• About 20% of the microorganisms posses EH genes
• The first epoxide hydrolase from A. niger (AnEH) was first
described in 1999
• identified in various bacteria, yeasts, and fungi
• including A. radiobacter, A. niger, M. tuberculosis, B. megaterium,
S. antibioticus, Pseudomonas spp., Corynobacterium spp.,
Norcardia spp., Arthrobacter spp., etc.
• Used for the conversion of many industrial epoxide
• Synthetic application like intermediates for pharmaceuticals drugs
like anti-obesity drugs, anti-depression drugs, HIV protease
inhibitors etc.

6. Structure
• The first epoxide structure published was of a bacterial
EH have sequence similarity with the mammalian EHs
• Divergent evolution
• all enzyme have common ancestor
but different catalytic properties
• Enzymes have a subunit mass of
35 to 40 kDa

7. Functions
• Microbes EHs involved in the
• Detoxifications of xenobiotic compounds,
• Metabolism of limonene for carbon natural source as well as
for intermediates, important for pharmaceuticals drugs like
Perillyl alcohol, Carvone, Terpineol
• Fungal EHs used for the production of mycotoxin
• Used for the synthesis of pure epoxide and diols
• Treatment of cystic fibrosis
• Treatment of Tuberculosis (lungs infection)

8. Detoxifications of Xenobiotic compounds

9. Synthetic Xenobiotic
• Some xenobiotics are resistant to degradation.
• They may be synthetic organochlorides such as plastics and
pesticides, or naturally occurring organic chemicals such as
polyaromatic hydrocarbons (PAHs) and some fractions of crude oil
and coal
• microorganisms epoxide hydrolases are capable of degrading
almost all the different complex and resistant xenobiotics found on
the earth than mammals EHs
• Aspergillus niger, White rot fungi (Phanerochaete chrysosporium)
• Mycobacterium, pseudomonas, Rhodococcus spp.
• Aspergillus, Penicillium , Flavobacterium
• E.coli

10. Metabolism of limonene
• In 1998, a bacterial (Rhodococcus erythropolis) epoxide
hydrolase used degradation pathway for limonene
• Limonene (4-isopropenyl-1-methylcyclohexene), a monocyclic
monoterpene, and is formed by more than 300 plants
• Most of the microbes hydrolyzed it with limonene 1,2-epoxide
hydrolase and used it as the source of carbon ( as engery)
• As well as

11. • Limonene is used to
synthesize
1. Perillyl alcohol
 used as an ingredient in
cleaning products and
cosmetics, also as a potential
treatment for people with
brain cancer
2. Carvone
 prevent premature sprouting
of potatoes during storage,
mosquito repellent, pesticide
3. Terpineol
 antibacterial and antiviral, an
immune system stimulant

12. Treatment of Tuberculosis (lungs infection)
• Tuberculosis is the major cause of death in HIV-infected
individuals caused by Mycobacterium tuberculosis .
• Two most important anti-tubercular agents, isoniazid and
rifampicin increased the resurgence of the disease
• The genome of Mycobacterium codes for large number of
potential epoxide hydrolases that converts the epoxide to diols
• The unusual number of EHs in the bacterium, combined with
the potential toxicity of EH substrates in general, suggests a vital
function for this enzyme family in the physiology of the
pathogen
• So blocking of EH function or inhibtors of EH may represent a
promising new approach for antitubercular therapy

13. Treatment of cystic fibrosis
• Cystic fibrosis (CF)
• is a disorder , affects mostly the lungs, pancreas, liver, kidneys, and intestine nearly
• 80% of patients with cystic fibrosis have a chronic infection due to P.aeruginosa (
bacteria)
• Treatment is difficult due to the formation of antibiotic-resistant biofilms by bacteria
• The bacteria decrease the efflux of chloride ion from lungs by disturbing the cystic
fibrosis ion channel (CFTR) in epithelial cell make it more suitable for its growth
• The downregulation of plasma membrane CFTR is mediated by a single secreted
protein, the CFTR inhibitory factor (Cif), Within an hour after treatment with Cif, the
levels of CFTR in the apical membrane are significantly reduce
• Based on sequence comparisons, Cif showed the greatest degree of sequence
similarity to the class of epoxide hydrolases (EHs)

14. • Verenium Corporation , located in San Diego, California,
constructed libraries of DNA isolated around the world. By
using activity-based high throughput assays they discovered
novel microbial EHs (about 50 In total). They are unique at
sequence as well as catalyctic activity levels.
• The work on epoxide hydrolases get important due to its
various role in mammals, plants, insects as well as in
microbes.
• So for many beneficial pharmaceuticals products the study
on the structure and function of epoxide in detail is
necessary.

15. References
• Morisseau, C. (2013). Role of epoxide hydrolases in lipid
metabolism. Biochimie, 95(1), 91-95.
• van der Werf, M. J., Overkamp, K. M., & de Bont, J. A. (1998).
Limonene-1, 2-epoxide hydrolase fromRhodococcus erythropolis
DCL14 belongs to a novel class of epoxide hydrolases. Journal of
bacteriology, 180(19), 5052-5057.
• Naworyta, A. (2010). Structure-function studies of epoxide
hydrolases (Vol. 2010, No. 5).
• van der Werf, M. J., Swarts, H. J., & de Bont, J. A. (1999).
Rhodococcus erythropolis DCL14 contains a novel degradation
pathway for limonene. Applied and environmental microbiology,
65(5), 2092-2102.

16. • Arand, M., Cronin, A., Oesch, F., Mowbray, S. L., & Alwyn Jones,
T. (2003). The telltale structures of epoxide hydrolases. Drug
metabolism reviews, 35(4), 365-383.
• Bahl, C. D., Morisseau, C., Bomberger, J. M., Stanton, B. A.,
Hammock, B. D., O'Toole, G. A., & Madden, D. R. (2010). Crystal
structure of the cystic fibrosis transmembrane conductance
regulator inhibitory factor Cif reveals novel active-site features
of an epoxide hydrolase virulence factor. Journal of
bacteriology, 192(7), 1785-1795.
• Orru, R. V., & Faber, K. (1999). Stereoselectivities of microbial
epoxide hydrolases. Current opinion in chemical biology, 3(1),
16-21.

17. • Johansson, P., Unge, T., Cronin, A., Arand, M., Bergfors, T.,
Jones, T. A., & Mowbray, S. L. (2005). Structure of an atypical
epoxide hydrolase from Mycobacterium tuberculosis gives
insights into its function. Journal of molecular biology, 351(5),
1048-1056.
• Fretland, A. J., & Omiecinski, C. J. (2000). Epoxide hydrolases:
biochemistry and molecular biology. Chemico-biological
interactions, 129(1), 41-59.
• Steinreiber, A., & Faber, K. (2001). Microbial epoxide
hydrolases for preparative biotransformations. Current opinion
in Biotechnology, 12(6), 552-558.

18. • Duetz, W. A., Bouwmeester, H., Van Beilen, J. B., & Witholt, B.
(2003). Biotransformation of limonene by bacteria, fungi, yeasts,
and plants. Applied microbiology and biotechnology, 61(4), 269-
277.
• Rink, R., Kingma, J., Lutje Spelberg, J. H., & Janssen, D. B. (2000).
Tyrosine residues serve as proton donor in the catalytic mechanism
of epoxide hydrolase from Agrobacterium radiobacter.
Biochemistry, 39(18), 5600-5613.
• Morisseau, C., Ward, B. L., Gilchrist, D. G., & Hammock, B. D.
(1999). Multiple epoxide hydrolases in Alternaria alternata f. sp.
lycopersici and their relationship to medium composition and host-
specific toxin production. Applied and environmental microbiology,
65(6), 2388-2395.

19. • Johansson, P., Unge, T., Cronin, A., Arand, M., Bergfors, T., Jones, T.
A., & Mowbray, S. L. (2005). Structure of an atypical epoxide
hydrolase from Mycobacterium tuberculosis gives insights into its
function. Journal of molecular biology, 351(5), 1048-1056.
• Gomez, G. A., Morisseau, C., Hammock, B. D., & Christianson, D. W.
(2004). Structure of Human Epoxide Hydrolase Reveals Mechanistic
Inferences on Bifunctional Catalysis in Epoxide and Phosphate Ester
Hydrolysis†. Biochemistry, 43(16), 4716-4723.
• Arand, M., Hallberg, B. M., Zou, J., Bergfors, T., Oesch, F., van der
Werf, M. J., ... & Mowbray, S. L. (2003). Structure of Rhodococcus
erythropolis limonene‐1, 2‐epoxide hydrolase reveals a novel active
site. The EMBO journal, 22(11), 2583-2592.