we wanted to see if different [substrate] affected velocity of rxn with an enzyme
We kept the enzyme volume always 0.2 mL optimal pH is 8. We did everything at pH 7 and total volume in cuvet was 3mL 0.2 mL enzyme + 0.3 mL substrate + 2.5 mL tris = total volume always 3ml
0.2 mL enzyme + 0.9 mL substrated + 1.9 mL tris = total volume always 3ml
We would put these cuvets under a spectrophotometer determined %T and graphed it.
Another group did the exact same thing so that we could compare our results to make sure they were similar.
But the group made an error. Instead of a total volume of 3mL they used a total volume of 5 mL
i.e. 0.2 mL enzyme + 0.3 mL substrate + 4.5 mL tris = total volume always 5ml
0.2 mL enzyme + 0.9 mL substrated + 3.9 mL tris = total volume always 5ml
They added more Tris. Now their %T values (at 5mL total) are higher than our %T values (at 3ml) so they have a lower [ ] when I used beers law to determine the [ ]. this lower [ ] is really skewing my data.
information i have:
Enzyme = 0.135g/700 mL optimal pH is 8
Tris buffer = 0.05M
Substrate = 1mM
Can I do anything to fix this?
They numbers are really skewing my data. I would prefer not to exclude their data. Any suggestions?