Uncompetitive inhibition
An uncompetitive inhibitor interacts with the enzymesubstrate complex, but not with the enzyme alone. For uncompetitive inhibition, the doublereciprocal equation is as follows:
where
This equation shows that a doublereciprocal plot of enzyme kinetic data with varying concentrations of an uncompetitive inhibitor should give a straight line with varying yintercepts, but with the same slopes (Figure 1). In enzyme kinetic assays with an uncompetitive inhibitor, the apparent
Figure 1: a) LineweaverBurk plot showing the uncompetitive inhibition. b) yintercepts of each linear regression plotted against the inhibitor concentration.
Calculating K’I, K_{m} and V_{max} K I, K m, and V max
When working with an uncompetitive inhibitor, no parameters can be calculated from the initial LineweaverBurk plots. To calculate the different parameters, the "yintercepts" of these plots must be plotted against the inhibitor concentration:
This plot should, therefore, result in a straight line with the intercept
Steps of calculating the kinetic parameters when using an uncompetitive inhibitor

Prepare LineweaverBurk plots of the kinetic data and fit the data using linear regression (1 fit per inhibitor concentration).

Plot the yintercepts of each of these fits as a function of the inhibitor concentration.

To calculate
V_{max}, V max take the reciprocal of the yintercept of this plot. 
To calculate
K_{I}, K I divide the yintercept of this plot with the slope. 
To calculate
K_{m}, go back to the first LineweaverBurk plots, take the slope from 1 of these plots (the slope should be the same, or close, for all the plots), and multiply it with V_{max}. K m, go back to the first LineweaverBurk plots, take the slope from 1 of these plots since the slope should be the same, or close, for all the plots, and multiply it with V max.
References
 Lehninger, Albert L.; Nelson, David L.; Cox, Michael M. (2008). Principles of Biochemistry (5th ed.). New York, NY: W.H. Freeman and Company. ISBN 9780716771081.