Help with Kinetics-double reciprocal plot

[skyblu280]

Hi,

I have an assignment where we basically have to make a Velocity vs. Substrate plot and find the Km and Vmax values.

We then have to do a double reciprocal plot and find the Km and Vmax from that.

My first question is if I have to convert the units of my axes so they are uniform.
The substrate is in micromolar while the Velocity is in micromolar/min. I asked my TA and he said no...but I just wanted to double check.

On the double reciprocal plot, I plotted the points on deltagraph and they gave me a straight line for the double reciprocal plot in the form of y=mx+b

Is KM the same as Ks in this case? I was thinking that I could get the value of Km but having the slope equal to Ks/Vmax. I can read Vmax off the graph and solve for Ks which I would determine it to be equal to KM. Is that a correct assumption?

 

[epenguin]

You use a lot of words which you do not define so it is difficult to answer.

For the question about 'uniformization' I guessed what you mean. If you have a table in two columns of 1/s, 1/v it should be obvious a reasonable way to plot them and you seem to have done that.

Slightly worrying loosenesses of expression. It is scarcely more difficult to say 'I can read 1/V_max from the intercept on the 1/v axis' in which case we would know you know what you are doing than it is to say vaguely 'I can read Vmax off the graph' in which case you may do but we are not sure.

You don't have to solve any equations to get your K, -1/K is the intercept on the 1/s axis, this must be in your textbook or lab manual. However what you say about the slope is also true, helps tie things up together.

I understand your question about K_m or K_s to mean 'is my measured constant practically an equilibrium constant, call it k_back/k_forward or is it a steady-state ('Briggs-Haldane') constant (k_back + k_cat)/k_forward, i.e. do I have a steady state removed from equilibrium?'

The answer is that as both cases give an equation of exactly the same form v = V_maxs/(K_m + s) the experiment gives you no information about this - you cannot tell the difference.

When you hand in your assignment do not forget to put units on your axis and any answers otherwise you will lose marks and more important, not help yourself in future understanding.