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Dear Ray:
Your dimer can sit on a crystallographic 2-fold and thus occupy the same
(perhaps even smaller) volume than a hairpin.
Most of the commercially available screens (including one based on a
screen I published in 1995, from Hampton) avoid citrate because it is
a chelating agent, and since the anion is a spectator that is presumably
repelled by the phosphori, my guess is that you could replace citrate
with any similar trivalent anion (or maybe 3x NaCl). The cation is
presumably what is important, and we have polluted the pdb and journals
with numerous hammerhead ribozymes crystallized in 3.6 M Li+ (1.8 M
Li2SO4). These crystals also grew in Na+ but the Li+ were better.
Our new full-length hammerhead ribozyme crystallized in NH4+ and PEG, but
the PEG phase-separates, so I am pretty sure the NH4+ ion is the relevant
ppt agent. I can't recall the details before my first coffee IV, but it
is listed in the paper (27 July 2006 Cell).
HTH,
Bill
On Thu, 17 Aug 2006 [EMAIL PROTECTED] wrote:
Hi folks,
Thanks for your interesting comments about RNA crystallization.
Actually I am trying to crystallize the RNA duplex, but the small amount of RNA
hairpin present in the sample appears to have come out, with the 1.4M
tri-sodium citrate.
The RNA has a self-complemantary sequence of course.
The crystals are quite dense and very birefringent and diffract to 2.8
angstroms. The small unit cell suggests that only a hairpin could fit in it.
I could not find any published report of RNA crystals coming out of tri-sodium
citrate. There are no divalent cations or polyamines present.
I am very surprised at this crystallization result.
Cheers,
Ray Brown
Department of Molecular and Cell Biology
University of Connecticut
________________________________
From: Daniel Anderson [mailto:[EMAIL PROTECTED]
Sent: Wed 8/16/2006 8:46 PM
To: Brown,Raymond (BIDMC - Experimental Medicine)
Cc: CCP4 News Group
Subject: Re: [ccp4bb]: RNA crystals
There's a nice article about RNA crystallization:
Golden, Kundrot (2003) Journal of Structural Biology 142, 98-107.
In a fast scan, I don't see mention of citrate ion.
It's amazing what I find in these piles around my desk.
-Dan
On Wed, 16 Aug 2006, William Scott wrote:
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It is fairly common to get RNA (or DNA) crystals in high concentrations of
monovalent cations, including Na+, Li+, and even NH4+. More surprising to
us was that several of the ribozymes function quite happily in the
presence of monovalent cations that were thought to require divalent
cations for cleaving.
One thing you will want to be aware of is that simple RNA hairpins have a
high propensity to crystallize as dimeric duplexes with mismatches in the
middle corresponding to the hairpin region.
The anion is usually not relevant, but in the case of citrate, it can
chelate Mg++. For that reason, I tend not to use it in screens, but there
is no particularly fundamental reason why citrate should be unusual, apart
from that.
HTH,
Bill
On Wed, 16 Aug 2006 [EMAIL PROTECTED] wrote:
Hi folks,
I appear to have grown some crystals of an RNA hairpin using 1.4 M sodium
citrate.
Has anybody else out there been able to get crystals of RNA or DNA using sodium
citrate as the precipitant ?
This result seems to be rather unusual to me.
Please get in touch if you know of any similar, successful crystallization
conditions.
Cheers,
Ray
email [EMAIL PROTECTED]
