kristoff at genbank.bio.net
Thu Mar 26 12:13:03 EST 1992
DRM21 at MB1.BIO.CAM.AC.UK (David Micklem) writes:
> I am making protein extracts from fruit fly ovaries and need to
>cause depolymerisation of the microtubules. Does anyone know whether this
>is possible to do by changing the buffer conditions, eg by including GDP or
>metal ion chelators? Id like to avoid using particularly stringent
>conditions like high salt/detergent levels if possible.
>Many thanks for your help.
It's been a while since I worked in this field, but fairly "low"
levels of Ca++ (i.e., uM or less) should do the trick (I hope that
this is not "high" salt by your use of the term). Metal ion chelators
are usually present in most microtubule *polymerization* buffers if my
memory has not failed me yet. Adding GDP would help, but I'd bet that
smaller amounts of Ca++ would be more effective. Since you are making
protein extracts you might not want to do the following, but small
additions of alkaline phosphatase can also chew up GTP and cause
microtubules to disassemble. Not being familiar with your system,
however, I can not say what side effects any of these treatments will
have on what you are trying to accomplish.
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