I'm performing inhibition study, since during competitive inhibition LB plot showing line of intercept on Y-axix (1/Vmax) is almost same for all inhibitor concentration used; with different Km (-1/Km) values. During non-competitive I found easier to calculate percentage of inhibition bcoz of different Vmax against control. should I use different Km against control Km as calculation factor?
Hi there,
Whatever the inhibition mechanism if you want to calculate a percentage of inhibition, you need to compare the initial velocities of the enzyme observed in absence and in presence of the inhibitor (all the conditions except inhibitor concentration being strictly identical). The ratio between the two will give you the percentage. But this percentage is not relevant as it is strictly related to the conditions used whereas Ki is unique for the system formed from one enzyme, one substrate and one inhibitor.
Hi Ramnath, the formula to calculate the percent of inhibition is %i = ((alpha - 1)/(alpha + beta))*100, where alpha = 1 + I/Ki and beta = S/Km; I is the inhibitor concentration, S the substrate concentration and Km is Michaelis-Menten constant. As you can see, %i depends on I, S, Ki and Km. If you have all these values you can calculate the %i. Good luck.
As both Dominique and Pedro stated, the critical thing you want to know is Ki. What you suggested was to compare inhibited vs uninhibited Km. If you do that, the ratio gives you 1 + i/Ki so you can easily work out Ki. However, you have to realise that the actual percent inhibition could be anything from zero (when [S] is much larger than the inhibited Km value) up to a large number limited only by inhibitor solubility! When [S] is small (relative to inhibited Km) v for any given substrate concentration will be proportional to 1/Km, and Km will be (1 + i/Ki) times uninhibited Ki. So e.g. if i = 100 times Ki then the rate will be decreased by a factor of 101 (approx. 99% inhibition). So there is no fixed number for the percentage you are looking for.
You can easily calclculate the % inhibition as indicated above by our colleagues.but this percent as accurate as it can be depends strictly on the experimental conditions used. In contrast the Ki is a more informative parameter because it is a real dissociation constant of the inhibitor.
Dear Ramnath, I want to know how to calculate the enzymatic antioxidants activities in ug/mg of protein from the OD. How to make standard curve. I have the values of OD at specific wavelength for SOD, POD, CAT, etc. I also want to know how to convert the OD into ug/ml for Total soluble protein. I would be really very thankful if you please explain.
Can someone please let me know which reference the source of this equation. Thank you, Ghassan
the percent of inhibition is %i = ((alpha - 1)/(alpha + beta))*100, where alpha = 1 + I/Ki and beta = S/Km; I is the inhibitor concentration, S the substrate concentration and Km is Michaelis-Menten constant. As you can see, %i depends on I, S, Ki and Km.
i need to calculate IC50 of extract using Enzyme inhibition. We are using substrate that is UV inactive. So we are using BSA to measure the quantity of the product. So we have absorbance and Fluoroscence intensity of Enzyme binding to the substrate and then we have taken OD and FI when in the same medium we added fixed concentration of BSA to obtain the amount of product formed. How will i calculate the IC50? We can calculate IC50 if we perform the kinetics of the drug, but it is applicable to pure compounds.
- Badges
- Science topic
- Similar topics
- Correction