[RASMB] AUC of RNA and RNA-proteins complexes

[Jo Butler]

Dear Pascal,

A v-bar of 0.55ml/g is more typical of DNA than RNA and you might well do 
better using 0.5ml/g for RNA.
If your synthetic nucleotides fold, as most RNA sequences do, 3S is not 
that surprising - tRNAs (70-80 nucleotides) are around 4S.

I have used AUC with both RNA and DNA complexes and life was not hell.  The 
main thing, particularly with RNA, is to be really pedantic about 
cleanliness of your cells to avoid any nuclease contamination.  The same, 
of course applies if you are working with protease sensitive proteins.
By following this principle, I have even run long-column equilibrium runs 
with RNA and, on other occasions, histones with not problem of degradation. 
Since the solutions were not sterilised, I can only assume that the 
contaminating bacteria were immobilised on the bottom of the cells and 
unable to secrete proteases or nucleases.

Jo Butler

--On Wednesday, March 6, 2002 5:06 pm -0800 pascal egea 
<pascal at msg.ucsf.edu> wrote:

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> Greetings,
> I am trying to do some AUC on a small synthetic RNA fragment (49
> nucleotides) and its complex with proteins by looking first at its
> sedimentation coefficient by velocity sedimentation.
> Does anybody have some experience about that? I heard that AUC on nucleic
> acids is hell and I would like to know if there are some good reference
> papers about this specific type of macromolecular systems?
> My actual S is about 3 S assuming a 0.55 ml/g V-bar (it seems very huge to
> me)...?
> what are the v-bar for RNA molecules? and what reasonnable value of S
> should I expect for a monomeric RNA piece of 49 nucleotide mainly folded
> as double stranded loop.
>
> Thank you in advance for your help
>
> Pascal Egea
> UCSF
> Department of Biophysics and Biochemistry
>
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P.J.G. Butler,
MRC Laboratory of Molecular Biology,
Hills Road, Cambridge, CB2 2QH, UK.
Tel. +44 (0)1223 402296