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Crystallization screens and methods: Difference between revisions
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Searching for silver bullets: An alternative strategy for crystallizing macromolecules. Alexander McPherson and Bob Cudney. Journal of Structural Biology 156 (2006) 387–406. | Searching for silver bullets: An alternative strategy for crystallizing macromolecules. Alexander McPherson and Bob Cudney. Journal of Structural Biology 156 (2006) 387–406. | ||
== Desiccation of an existing screen which shows no sign of crystallization or precipitation == | |||
''This reports a discussion on CCP4BB started on April 7, 2008 under the topic "which concentrated salt has lowest vapour pressure?".'' | |||
If, after setup of a crystallization experiment and waiting long enough, the droplets stay clear, the question arises whether one can modify this experiment to learn something. | |||
Thus the idea is that instead of setting up a new experiment, one simply modifies the reservoir in an attempt to make the protein crystallize (or precipitate), with the goal of bracketing the range of precipitant concentration. | |||
This idea has been explored in a number of publications: | |||
* H. Schreuder et al. published a method to measure the dehydration effect of precipitants (J. Appl. Cryst. (1988). 21, 426-429.). This publication has tables of the chemical potential of water in solutions of MPD, NaCl, potassium tratrate and PEG 6K. | |||
* similar experiments were done with NaCl, AS and MgSO4 by Arakali et al (Acta D 51 772-779, 2005) | |||
* AmSO4, NaCl and PEGs were explored in J. Newman, [doi:10.1107/S0907444905002726] Expanding screening space through the use of alternative reservoirs in vapor-diffusion experiments. Acta Cryst. (2005). D61, 490-493 | |||
* AmSO4 didn't work well; the ammonia changes the pH: Dunlop & Hazes, Acta Cryst D61, 1041-8, (2005) | |||
* Thakur AS, Robin G, Guncar G, Saunders NF, Newman J, Martin JL, Kobe B. [http://www.ncbi.nlm.nih.gov/pubmed/17971854?ordinalpos=3&itool=EntrezSystem2.PEntrez.Pubmed.Pubmed_ResultsPanel.Pubmed_RVDocSum] Improved success of sparse matrix protein crystallization screening with heterogeneous nucleating agents. PLoS ONE. 2007 Oct 31;2(10):e1091. They used dried seaweed; horse hair; cellulose and hydroxyapatite. | |||
further findings/ideas: | |||
* LiCl seems to work very well in the reservoir, whereas in the droplet Li salts tend to increase the solubility of peptides (Seebach et al., | |||
Helv. Chim. Acta 72 (1989) 857-867) - but on the other hand Li salts can also be used as cryoprotectants. | |||
* ~0.5 M LiCl match ~40% PEG4000 in chemical potential, and ~1.5-2 M LiCl match ~60-70% ammonium sulfate (~2.4 - 3 M). | |||
* at least 50% of PEG 4K, 8K, 10K desiccate drops. At less concentration PEGs seem to be poor. | |||
* add 100% glycerol or ethylene glycol | |||
* a number of substances [like dryrite, calcium sulfate (the salt that is used to absorb the humidity in damp rooms), and the gel-like stuff that keeps diapers dry] were mentioned, but they seem less appropriate to the author, for different reasons. | |||