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Heat extraction from a crystal depends on the thermal conductivity AND
the temperature gradient. If you start with a colder crystal you can
tolerate a larger temperature gradient before hitting the glass
transition temperature, and thus, in principle, transfer heat more
rapidly. I believe initial tests with Helium cooling didn't end up
helping much if anything compared to lN2 because Helium has a poor heat
capacity compared to N2 gas. I think these initial tests were reported
by Gert Rosenbaum and others but I may be wrong here.
Bart
Ethan Merritt wrote:
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Ian wrote>
Unless of course the crystal isn't as cold as you think it is!
Essentially all of the energy of absorbed X-ray photons must ultimately
be degraded to thermal energy, and unless this is efficiently conducted
to the surface where it can be removed by the cold stream, it's going to
produce some 'hot spots' which given sufficient X-ray flux may be
sufficient to cause local melting of the water glass.
On Tuesday 25 April 2006 09:05 am, Sebastiaan Werten wrote:
In which case one might expect better stability of crystals if helium
cooling is applied (ceteris paribus). Has this been observed?
If you mean "can a colder crystal withstand a more intense beam?"
the "expected" improvement has not, to my knowledge, been observed.
Some references here:
http://biop.ox.ac.uk/www/lj2000/garman/garman_01.html
My understanding is that the limiting factor is the thermal conductivity
of the crystal itself. Heat (photons) is deposited internally, and
must be transferred through the body of the crystal and possibly through
a bead of surrounding frozen buffer before it can be lost to the
external cooling stream. Making the crystal colder does not by itself
make this internal transfer any faster unless it also changes the
thermal conductivity. So if you start with a colder crystal it may
take fractionally longer to reach a harmful temperature, but the
x-ray flux that can be handled at equilibrium without continued
temperature rise remains the same.
One could speculate that the thermal conductivity will increase at
sufficiently cold temperatures (superconductivity), but I am dubious
this is relevant to protein crystals.
Note that this is a slightly different question than "does the crystal
diffract better, or decay more slowly". The latter is a question that
can be asked of a crystal in thermal equilibrium at various temperatures.
Hanson et al, J. Synchrotron Rad. (2002). 9, 375-381.
Perhaps someone can contribute additional pointers to relevant work.
Sebastiaan Werten.
----------
Unless of course the crystal isn't as cold as you think it is!
Essentially all of the energy of absorbed X-ray photons must ultimately
be degraded to thermal energy, and unless this is efficiently conducted
to the surface where it can be removed by the cold stream, it's going to
produce some 'hot spots' which given sufficient X-ray flux may be
sufficient to cause local melting of the water glass. There could well
be a steep temperature gradient, liq N2 temperature at the surface and
much warmer towards the centre. Also if low molecular weight solutes
are present the melting point could be locally depressed way below that
of pure water, so there wouldn't be quite so far to go to cause melting.
Admittedly I haven't looked at any calculations of what order of
magnitude the heating and heat conduction effects might be for a typical
flux and I could be way off beam but it's a least a theoretical
possibility.
-- Ian
-----Original Message-----
From: [EMAIL PROTECTED] [mailto:[EMAIL PROTECTED] On
Behalf Of James Holton
Sent: Tuesday, April 25, 2006 3:11 PM
To: Udupi Ramagopal
Cc: Uhnsoo Cho; [EMAIL PROTECTED]
Subject: Re: [ccp4bb]: Radiation damage problem
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It is definitely true that some sensitive residues decay quite a bit
faster than the high-resolution spots. This has been repoted
in several
papers now. What has not been repoted (to my knowledge) is a
convincing
connection between the decay of side chains involved in a crystal
contact and the loss of high-resolution data. In all of the
studies I
have seen that measure the rate of loss of resolution, it is always
tightly related to dose. If you know of a study where it has
been shown
to be related to something else, I would like to hear about it.
I, personally, do not think that there is a connection between
specific damage near crystal contacts and loss of resolution. My
reasoning is that a cryo-cooled crystal is permeated by a matrix of
glassy water. Glassy water is a SOLID. So, once cooled, the crystal
lattice is supported by a lot more than just crystal
contacts. Kind of
like an insect preserved in amber. Breaking bonds near the
contacts are
not going to do much more distortion to the overall lattice
than broken
bonds in the core or even in the solvent. Diffraction is a
process that
occurs over dozens of unit cells. There must be long-range
distortions
to impact it. Site-specific damage, formally, only impacts
the B-factor
of that site.
-James Holton
MAD Scientist
Udupi Ramagopal wrote:
I think some times the sensitivity of residues involved in critical
cystal contacts
can disturb these valuable suggestions and calculations.
Ramu
On Mon, 24 Apr 2006, James Holton wrote:
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Disclaimer:
Radiation damage is a complex phenomenon, and many aspects
of radiation
damage at low temperature are still hard to explain, let
alone predict.
You DEFINITELY need to know things like the beamline's
brightness and
the concentrations of any atoms in your sample heavier
than oxygen to
even have a ghost of a chance of understanding the origin of your
problem.
However...
If you are used to this beamline and you "feel" that these
crystals are
decaying faster than "normal", then the most likely
problem is that you
have a heavy atom in your sample. This will cause your
crystal to absorb
a lot more x-rays than normal (doing more damage).
Anything heavier than
oxygen is a potential problem, and stuff in the solvent counts! The
"rule of thumb" is that the "absorptivity" of an atom is roughly
proportional to the atomic number. So, try doing things
like replacing
potassium with lithium or iodide with fluoride (each a
factor of 6).
Obviously, the more concentrated a heavy atom is, the
bigger the problem
will be.
Since beamline brigthnesses vary by a factor of 30,000
from place to
place, and crystal absorptivity can vary by a factor of 10 or more,
radiation damage questions can be hard to answer without
knowing flux,
beam size and sample composition.
But, if you do know these things...
The decay in protien crystals is generally proportional to
absorbed dose
(Joules/kg or "Gray"). Dose, in turn is proportional to fluence
(photons/mm^2) and the "extinction coefficient" of your
crystal in the
x-ray range. You get fluence (photons/mm^2) from flux
(photons/s) by
dividing by the beam size to get brightness (photons/mm^2/s) and
multiplying by the total exposure time (in seconds). Given
the beamline
flux, wavelength, beam size, and crystal composition the expected
lifetime of your crystal can be computed with a program
called RADDOSE
(Murray et. al. JSR (2005), 12, 268-75). Or you can "google" for
RADDOSE.
For example, you can expect the lifetime of your crystal will be
(roughly) cut in half by the addition of 4M NaCl, 5M
NH4SO4, 1 Se per 30
residues (at 0.97A), or 150 mM CsI.
-James Holton
MAD Scientist
Uhnsoo Cho wrote:
Dear bulletin members,
Sorry for non-CCP4 related questions.
Recently, I've got crystals which have a radiation damage problem.
It also has a weak diffraction because of the big unit
cell dimension
(about 300A in one direction), so I have to overexpose
crystals even
with the synchrotron beam.
The problem is that during the data collection, the
resolution decaded
gradually from 3 to 6A after certain number of frames
(about 30 or 40
frames).
Do you have any experiences or any suggestions that I can
evade this
radiation damage?
Any suggestions will be grateful.
Thank you.
Best regards,
~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~
Uhn-Soo,Cho
Graduate student in Biological Structure Department
University of Washington.
HSB G514
1959 NE Pacific St.
Seattle, WA 98195-7420
Box. 357420
Fax #206-543-1524
Tel # 206-221-2435
~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~
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