Km Vmax - (Mar/22/2013 )
I have few questions on this matter, let me give one example that is 'the reported Km value of glucose for GLUT1 in human is 10mM'.
My questions are:
1) Why do we need to refer to Km value and used that enzyme concentration in enzyme/transporter related work?
2) Will this glucose Km value be the same for all people or it varied?
3) Will this value vary across species ie human vs rat?
4) Will we get the same Km value in in vitro and in vivo experiment for GLUT1?
5) Can we simply take reported Km value and use it for experiment or it will be best for determining for each new experiment?
Looking forward to hear from you guys.
Much of my work is on enzyme kinetics. Km values will differ from different labs and definetly in different species. So much of the Km value is based on the purity of the enzyme in the assay. If you have impure and/or dead enzyme the Km value will likely be off. Buffer conditions greatly affect Km values. Here is a great webpage on developing enzymatic assays:
The Km value is important because the concentrations you use in assays need to cover a range surrounding the Km value.
Question 4: in vitro vs in vivo Km values. In theory, if the conditions in vitro are exactly the same, the km values should be very close. However, inhibitors could be present in vivo, that could inflate the apparent Km value.
You should always try to replicate Km data. You should try to be in the same range as previously reported literature, unless you have optimized the assay and you now have superior (lower) Km values.
The better kinetic parameter to care about is overall efficiency (kcat/Km). This is much more insightful compared to a Km value alone.