Different Plots for detemination pf Km and Vmax

Different plots for Determination of km and vmax

1. Different Plots for Determination of Km and Vmax SUBMITTED BY: Amir sohil SUBMITTED TO: Bharat Bhusan Sir

2. Vmax Vmax is the reaction rate when the enzyme is fully saturated by substrate, indicating that all the binding sites are being constantly reoccupied . Km Km is a value of substrate concentration at half maximal velocity

3. Michaelis-Menten Equation Michaelis –Mentum kinetics is one of the simplest and best known model of enzyme kinetics. This model explain how an enzyme can cause kinetic rate enhancement of a reaction and why the rate of reaction depends on the concentration of enzyme present.

4. According to this model the enzyme reversibly bind with substrate to form an ES complex that subsequently yield product.

5. The mechanism of enzyme catalyzed reactions is studied by making kinetic measurements on enzyme substrate reaction system The simple kinetic model os obtained, when plotting the rate of catalysis,V (reaction velocity),V/S the substrate concentration [S]

6. From this equation, the reaction velocity,V can be expressed as .

7. Lineweaver-Burk Plot The Lineweaver-Burk plot is a graphical representation of the Lineweaver-Burk Equation of Enzyme kinetics,described by Lineweaver and Dean Burk in 1934.

8. Taking the reciprocal of this equation gives

9. This plot was widely used to determine Km and Vmax, before the wide availability of powerful computers and non-linear regression Software.

10. Eadie-Hofstee and Hanes Plot The Lineweaver-Burk plot has been criticized on several grounds Firstly the extrapolation across the 1/Vo axis to determine _1/Km sometimes reaches the edge of graph paper before reaching 1/[S] axis Secondly, it is said to give undue weight to measurements made at low concentration.

11. Eadie-Hofstee Plot

12. The Eadie-Hofstee plot takes as its starting point the Lineweaver-Burk equation.Both sides of the equation are multiplied by the factor Vo V max :

13. The Hanes plot (devised by Charles Hanes) similarly starts with the Lineweaver-Burk equationwhich in this instance is multiplied throughout by [S]

14. Eisenthal and Cornis Bowden Plot In view of the difficulties in obtaining reliable estimates of Km and V max , even with the aid of statistical analysis, from the plots discussed above, Robert Eisenthal and Athel Comish-Bowden (1974) suggested a different approach, though one still based on the Michaelis-Menten equation (7.12). The reciprocal form of this equation, at constant [E0], gives

15. Therefore, at constant v0 and [S0], a plot of V max against Km is linear.

16. References Palmer Trevor,Bonnner Philip Enzymes second edition Gupta N.S Biochemistry