Kinetics of enzyme activity SlideShare

Kinetics of enzyme activity SlideShare

1. Kinetics of enzyme activity By Sayanta Mitra

2. CONTEXT Enzyme Classification of enzymes Structure of enzymes Fischer’s Lock and Key Model Michaelis-Menten Model Factors affecting enzyme activity Enzyme inhibition Conclusion

3. What is Enzyme? Proteins that act as biological catalysts . Increase the rate of a reaction without being used up or changed themselves. Enzymes act upon the molecules are known as substrates . Coenzyme : A substance that enhances the action of an enzyme . They cannot by themselves catalyze a reaction but they can help enzymes to do so. Coenzymes are organic non-protein molecules that bind with the protein molecule (apoenzyme) to form the active enzyme (holoenzyme).

4. Apoenzyme : A protein that forms an active enzyme system by combination with a coenzyme and determines the specificity of this system for a substrate is called apoenzyme. Holoenzyme : Holoenzymes are the active forms of enzymes. Enzymes that require a cofactor but are not bound by one are called apoenzymes. Holoenzymes represent the apoenzyme bound to its necessary cofactors or prosthetic groups

5. Classification of enzymes : International Union of Biochemistry (I.U.B.) in 1964 has adopted classification of enzymes depending on the type of reactions they catalyze. According to this commission, all enzymes are classified into 6 major classes Oxidoreductases- Oxidation-reduction reactions (transfer of electrons). Transferases- Transfer of group Hydrolases- Hydrolytic reactions (transfer of functional groups to water) Lyases- Addition or removal of groups to form double bonds Isomerases- Transfer of groups within molecules to yield isomeric forms Ligases- Condensation of two molecules coupled through ATP hydrolysis

6. Enzyme Structure : Enzymes are proteins and usually have a globular tertiary structure. Enzyme has the active site . Active site : The active site of an enzyme is the region that binds the substrate and converts it into product . Three-dimensional structure. Consists of portions of a polypeptide chain . Enzyme binds its substrate and form enzyme–substrate complex.

7. Fischer’s Lock and Key Model : Model is proposed by Emil Fischer . The substrate can fit into its complementary site on the enzyme as a key fits into a lock . Two shapes are rigid and fixed . Perfectly complement each other.

8. Kinetics of Enzyme : Michaelis-Menten Model : The Michaelis-Menten model is one of the simplest and best-known approaches to enzyme kinetics. Michaelis-Menten kinetics is a model of enzyme kinetics which explains how the rate of an enzyme-catalysed reaction depends on the concentration of the enzyme and its substrate. Let’s consider a reaction in which a substrate (S) binds reversibly to an enzyme (E) to form an enzyme-substrate complex (ES), which then reacts irreversibly to form a product (P) and release the enzyme again. S + E ⇌ ES → P + E Two important terms within Michaelis-Menten kinetics are:

9. V max  – T he maximum rate of the reaction , when all the enzyme’s active sites are saturated with substrate. K m  – T he substrate concentration at which the reaction rate is  50% of the V max . Km is a measure of the affinity, an enzyme has for its substrate. As the lower the value of Km , the more efficient the enzyme is at carrying out its function at a lower substrate concentration . The Michaelis-Menten equation for the reaction is: v = Velocity of reaction
V max = Maximum rate achieved by the system
[S] = Concentration of a substrate S
K m = Michaelis constant

10. Factors affecting enzyme activity : Temperature : Temperature affects the rate of enzyme-catalyzed reactions in two ways,- First, a rise in temperature , increasing the thermal energy of the substrate molecules. This raises, proportion of substrate molecules with sufficient energy to overcome the Gibbs free energy of activation (ΔG‡) . Hence increases the rate of reaction. The difference in energy level between the substrates and products is termed the change in Gibbs free energy (ΔG). The difference in free energy between the substrate and the transition state is termed the Gibbs free energy of activation (ΔG‡). Second, Increasing the thermal energy of the molecules make up the protein structure of the enzyme itself will increase the chances of breaking the multiple weak, non-covalent interactions (hydrogen bonds, van der Waals forces, etc.) which hold the three-dimensional structure of the enzyme together Ultimately this will lead to the denaturation of the enzyme.

11. pH : Each enzyme has an optimum pH at which the rate of the reaction that it catalyzes is at its maximum. Small deviations in pH from the optimum value lead to decreased activity due to changes in the ionization of groups at the active site of the enzyme. Larger deviations in pH lead to the denaturation of the enzyme protein itself, due to interference with the many weak non-covalent bonds maintaining its three-dimensional structure. The effect of (a) temperature and (b) pH on enzyme activity

12. Enzyme inhibition : Substances which decrease the rate of an enzyme catalyzed reaction are called as enzyme inhibitors and the process is known as enzyme inhibition. There are three types of reversible inhibitions: Competitive inhibition - In competitive inhibition, there is close resemblance in the structure of inhibitor I and substrate S, therefore, they both compete for the same active site on the enzyme. Competitive inhibitors decreases the rate of reaction by reducing the amount of active enzyme molecules bound to a substrate. The enzyme can form enzyme-substrate ES complex or it can form enzyme-inhibitor EI complex but not both ESI. Non-Competitive inhibition - In this type of inhibition, inhibitor has no structural similarity with substrate and binds with the enzyme at different site other than the active site.

13. Competitive inhibitors decreases the rate of reaction by reducing the amount of active enzyme molecules bound to a substrate. At very high substrate concentration, the chances of binding of inhibitor molecule to the enzyme will be reduced, so Vmax for the reaction will not be changed. Uncompetitive inhibition- In this type of inhibition, inhibitor does not bind to free enzyme. It binds only to enzyme substrate (ES) complex, directly or its binding is facilitated by the conformational change that takes place after substrate binds to enzyme. In both the cases, the inhibitor does not compete with substrate for the same binding site. Therefore, the inhibition cannot be overcome by increasing substrate concentration. Both Km and Vmax values are altered.

14. Conclusion : Enzyme kinetics finds its usefulness in various reactions mediate by enzyme , which include biochemical reactions. Enzymes work best in their optimum condition . Enzyme are used in Foods and beverages processing, animal nutrition, textile, etc.