INTRODUCTION Enzyme catalysis Mechanisms of catalysis Organic solvent Classes of Organic Solvent Biocatalysis in organic solvents Enzyme Reactions in Organic solvents Effect of the organic solvent in enzyme catalysis Conclusion References

1. ENZYME CATALYSIS
EFFECT OF ORGANIC SOLVENT
By
KAUSHAL KUMAR SAHU
Assistant Professor (Ad Hoc)
Department of Biotechnology
Govt. Digvijay Autonomous P. G. College
Raj-Nandgaon ( C. G. )

2. SYNOPSIS
INTRODUCTION
Enzyme catalysis
Mechanisms of catalysis
Organic solvent
Classes of Organic Solvent
Biocatalysis in organic solvents
Enzyme Reactions in Organic solvents
Effect of the organic solvent in enzyme catalysis
Conclusion
References

3. Enzyme catalysis
Enzyme catalysis is the catalysis of
chemical reactions by specialized proteins
known as enzymes
By providing an alternative reaction route and
by stabilizing intermediates the enzyme
reduces the energy required to reach the
highest energy transition state of the reaction
The mechanism of enzyme catalysis is similar in
principle to other types of chemical catalysis.

4. Mechanisms of
catalysis
•Enzymes are very specific. Each enzyme will
catalyze only one type of reactions and often will
only work with a specific substrate.
Electrostatic catalysis
Acid-base catalysis
Covalent catalysis
Metal ion catalysis

5. The Active Site
 Enzymes are typically HUGE proteins, yet only a small
part are actually involved in reaction.
The active site has two
basic components.
catalytic site
binding site
Model of
trios-p-isomerase

6. Catalytic site
Where reaction
occurs
Binding
Site
holds
substrate
in
place Substrate
Enzyme
The Active Site

8. Lock and Key Theory
Enzyme is “lock” and Substrate is the “key”.
Substrate structure must fit into enzyme’s structure.

9. Induced Fit Theory
Active site may not fit substrate.
Site must change in order to form the complex.

10. Enzymes have optimal operating temperatures and pHs.
These graphs show how enzyme catalysis is affected by
changes in temperature and pH for enzymes that operate
most effectively at physiological conditions.
Temperature and pH Effects on
Enzyme Catalysis
10

11. V(V)
Cr(VI)
Mn(VII)

12. •Common Classes of Organic Solvents
Chlorinated Aliphatic hydrocarbons Aromatic
hydrocarbons
trichloroethane
trichloroethylene
methylene chloride
chloroform
carbon tetrachloride
tetrachlorethylene
n - hexane
n - heptane
pentane
cyclohexene
cyclohexane
petroleum ether
benzene
toluene
xylenes
naphthalenes
Alcohols/Glycols/Ethers Esters/Ketones/Aldehydes Others
methanol
ethanol
propanol
butanol
ethylene glycol
diethyl ether
ethyl acetate
acetone
methyl ethyl ketone
methyl isobutyl ketone
methyl n-butyl ketone
formaldehyde
glutaraldehyde
carbon disulphid
pyridine
amides
amines

13. Biocatalysis in organic solvents
•Systems with organic solvents
•Water and a water miscible
organic solvent
•Two-phase systems
•PEG-modified enzymes in
organic solvents
•Reversed micelles
•Monophasic organic solvents

14. EFFECT OF ORGANIC SOLVENT ON ENZYME CATALYSIS
Effects on enzyme stability
Hydrophobic solvent: small redistribution of water: conservation of
native protein structure
Polar solvent: stronger effect
Interaction of solvent with protein surface
Strip tightly bound water
Destruction of hydrogen bond network
Lowering of surface tension
Onset of protein unfolding
Extreme thermostability in inert solvents Fewer side reactions
(hydrolysis)
Conformational rigidity in dehydrated state
Ex.Chymotrypsin,

15. Water-water miscible organic solvents
Polar solvents
Reactant, inhibitor, increase of flexibility (rate)
Operational stability
Change in product pattern
Substrate solubility
Presence of organic solvent can have a
large effect on substrate solubility
A substrate with a low affinity for solvent
binds strongly to the enzyme
Polar substrates have high Km in polar
solvent

16. Two-phase systems
About equal volumes of an aqueous solution and an immiscible
organic solvent
Catalysis takes place in the aqueous phase or at the interface
[S] low, limits rate of catalysis
Traces of solvent can influence activity and stability
Enzyme recovery is difficult

17. PEG-modified enzymes
Modification of lysine residues with PEG molecules of different size
Synthesis of organic solvent soluble enzymes
10 - 20 PEG chains per enzyme molecule
Increase in molecular mass
Protects enzyme from surrounding organic solvent and prevents
stripping of essential water
High enzymatic activity with water immiscible solvents

18. Reversed micelles
Effects on enzyme stability
- dependent on protein properties
low water content increases stability
Effects on enzyme activity
Water content too low
pH different from stock buffer
solution due to binding of protons or
hydroxyl groups with surfactant head
groups
Effects on enzyme kinetics
effects substrates
Increase in apparent Km

19. Effect of solvent on the water associated with the
enzyme
water-stripping in the hydrophilic solvents
Since enzyme and solvent compete for water, optimal water
content is related to the amount of enzyme and the
concentration of substrate
hydrophilization of enzyme surface minimize removal of
the H2O from enzyme

20. Effect of solvent on the enzyme
Protein dynamics
Protein conformation
Enzyme active center
Effect of solvent on substrates and products
•change the conce. of substrates or products in the
aqueous layer around enzyme
Effect of substrate solvation on kinetics
increase Km app in organic solvent
solvent affects enzyme activity and specificity by altering the
partitioning of substrates and products

21. CONCLUSION
The technological utility of enzymes can be enhanced greatly by using them
in organic solvents rather than in their natural aqueous reaction media.
Strong acids and bases, organic solvents, mechanical
action, and high temperature decrease an enzyme-
catalyzed rate of reaction. Even slight changes in the pH
can have profound effects on enzyme catalysis

22. REFERENCES
•Principle of Biochemistry (fifth edition) by Nelson & Cox
•Biochemistry (fourth edition) by Donald voet and Judth G.
Voet
•Klijn, J and Engberts, J. Organic
chemistry: Fast reactions 'on
water'. Nature 435, 746-747 (9 June 2005)
•www.wikipedia.com