[RASMB] active enzyme centrifugation

[Lake Paul]

Judith,

A few things to take into consideration:
1. What is the smallest and largest possible species that might form? That
will determine your fastest rotor speed (e.g clearance of the optical path)
2. You can estimate the fastest speed using the equation published in
Schuck, P et. al  Basics of Analytical Ultracentrifugation.  The equation
is on Pg 213.
3. You will more than likely need to do a multi-speed SV experiment. This
can take a few forms, but I would just do a single concentration at two
speeds. Double the concentration and repeat. Another way would be to spin
at a low speed, collect data then speed up and collect data.
4. The reaction kinetics (fast and slow equilibrium) of the interaction
will also examined.

Those are some of the things I can think of off the top of my head. More
might come later.

Lake

Lake




On Thu, Apr 4, 2019 at 12:35 PM Judith Kornblatt <
Judith.Kornblatt at concordia.ca> wrote:

> >
> >
> >> I'm doing active enzyme centrifugation on an olligomeric enzyme,
> >> hoping to learn whether or not the individual subunits have activity.
> >> I know how to manually analyze the data when there is only one active
> >> species, but don't know how to do the analysis if there are multiple
> >> active species with different s values. In addition, I'd prefer to
> >> not have to do this manually. Any suggestions for the analysis?
> >>
> >> Judith Kornblatt
> >> Profesor Emeritus
> >> Department of Chemistry and Biochemistry
> >> Concordia University, Montreal
> >>
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