https://wiki.uni-konstanz.de/ccp4/index.php?title=Crystal_growth:_Protein-DNA_complexes&feed=atom&action=history Crystal growth: Protein-DNA complexes - Revision history 2024-03-28T20:27:33Z Revision history for this page on the wiki MediaWiki 1.39.6 https://wiki.uni-konstanz.de/ccp4/index.php?title=Crystal_growth:_Protein-DNA_complexes&diff=2342&oldid=prev Kay at 08:54, 9 September 2015 2015-09-09T08:54:11Z <p></p> <table style="background-color: #fff; color: #202122;" data-mw="interface"> <col class="diff-marker" /> <col class="diff-content" /> <col class="diff-marker" /> <col class="diff-content" /> <tr class="diff-title" lang="en"> <td colspan="2" style="background-color: #fff; color: #202122; text-align: center;">← Older revision</td> <td colspan="2" style="background-color: #fff; color: #202122; text-align: center;">Revision as of 09:54, 9 September 2015</td> </tr><tr><td colspan="2" class="diff-lineno" id="mw-diff-left-l16">Line 16:</td> <td colspan="2" class="diff-lineno">Line 16:</td></tr> <tr><td class="diff-marker"></td><td style="background-color: #f8f9fa; color: #202122; font-size: 88%; border-style: solid; border-width: 1px 1px 1px 4px; border-radius: 0.33em; border-color: #eaecf0; vertical-align: top; white-space: pre-wrap;"><div>7. Other useful tips?</div></td><td class="diff-marker"></td><td style="background-color: #f8f9fa; color: #202122; font-size: 88%; border-style: solid; border-width: 1px 1px 1px 4px; border-radius: 0.33em; border-color: #eaecf0; vertical-align: top; white-space: pre-wrap;"><div>7. Other useful tips?</div></td></tr> <tr><td class="diff-marker"></td><td style="background-color: #f8f9fa; color: #202122; font-size: 88%; border-style: solid; border-width: 1px 1px 1px 4px; border-radius: 0.33em; border-color: #eaecf0; vertical-align: top; white-space: pre-wrap;"><br/></td><td class="diff-marker"></td><td style="background-color: #f8f9fa; color: #202122; font-size: 88%; border-style: solid; border-width: 1px 1px 1px 4px; border-radius: 0.33em; border-color: #eaecf0; vertical-align: top; white-space: pre-wrap;"><br/></td></tr> <tr><td class="diff-marker" data-marker="−"></td><td style="color: #202122; font-size: 88%; border-style: solid; border-width: 1px 1px 1px 4px; border-radius: 0.33em; border-color: #ffe49c; vertical-align: top; white-space: pre-wrap;"><div>Protein DNA crystals can be very fragile so good idea to screen with cryoprotectant in the screen to save manipulations</div></td><td class="diff-marker" data-marker="+"></td><td style="color: #202122; font-size: 88%; border-style: solid; border-width: 1px 1px 1px 4px; border-radius: 0.33em; border-color: #a3d3ff; vertical-align: top; white-space: pre-wrap;"><div><ins style="font-weight: bold; text-decoration: none;">* </ins>Protein DNA crystals can be very fragile so good idea to screen with cryoprotectant in the screen to save manipulations</div></td></tr> <tr><td class="diff-marker" data-marker="−"></td><td style="color: #202122; font-size: 88%; border-style: solid; border-width: 1px 1px 1px 4px; border-radius: 0.33em; border-color: #ffe49c; vertical-align: top; white-space: pre-wrap;"><div>Test crystals with a gel to see if DNA is present</div></td><td class="diff-marker" data-marker="+"></td><td style="color: #202122; font-size: 88%; border-style: solid; border-width: 1px 1px 1px 4px; border-radius: 0.33em; border-color: #a3d3ff; vertical-align: top; white-space: pre-wrap;"><div><ins style="font-weight: bold; text-decoration: none;">* </ins>Test crystals with a gel to see if DNA is present</div></td></tr> <tr><td class="diff-marker" data-marker="−"></td><td style="color: #202122; font-size: 88%; border-style: solid; border-width: 1px 1px 1px 4px; border-radius: 0.33em; border-color: #ffe49c; vertical-align: top; white-space: pre-wrap;"><div>Use of SAXS to get molecular envelop of complex</div></td><td class="diff-marker" data-marker="+"></td><td style="color: #202122; font-size: 88%; border-style: solid; border-width: 1px 1px 1px 4px; border-radius: 0.33em; border-color: #a3d3ff; vertical-align: top; white-space: pre-wrap;"><div><ins style="font-weight: bold; text-decoration: none;">* </ins>Use of SAXS to get molecular envelop of complex</div></td></tr> <tr><td colspan="2" class="diff-side-deleted"></td><td class="diff-marker" data-marker="+"></td><td style="color: #202122; font-size: 88%; border-style: solid; border-width: 1px 1px 1px 4px; border-radius: 0.33em; border-color: #a3d3ff; vertical-align: top; white-space: pre-wrap;"><div><ins style="font-weight: bold; text-decoration: none;">* general guidelines to prepare protein/DNA complexes suitable for crystallization trials: [http://dx.doi.org/10.1007/978-1-59745-209-0_11 Hollis T (2007) Crystallization of protein-DNA complexes. Methods Mol. Biol. 363: 225–237]</ins></div></td></tr> <tr><td colspan="2" class="diff-side-deleted"></td><td class="diff-marker" data-marker="+"></td><td style="color: #202122; font-size: 88%; border-style: solid; border-width: 1px 1px 1px 4px; border-radius: 0.33em; border-color: #a3d3ff; vertical-align: top; white-space: pre-wrap;"><div><ins style="font-weight: bold; text-decoration: none;">* design and composition of a specialized screen for protein/DNA complexes: [http://dx.doi.org/10.1107/S1744309112025316 Pryor EE Jr, Wozniak DJ &amp; Hollis T (2012) Crystallization of Pseudomonas aeruginosa AmrZ protein: development of a comprehensive method for obtaining and optimization of protein-DNA crystals. Acta Crystallogr. Sect. F Struct. Biol. Cryst. Commun. 68: 985–993]</ins></div></td></tr> <tr><td class="diff-marker"></td><td style="background-color: #f8f9fa; color: #202122; font-size: 88%; border-style: solid; border-width: 1px 1px 1px 4px; border-radius: 0.33em; border-color: #eaecf0; vertical-align: top; white-space: pre-wrap;"><br/></td><td class="diff-marker"></td><td style="background-color: #f8f9fa; color: #202122; font-size: 88%; border-style: solid; border-width: 1px 1px 1px 4px; border-radius: 0.33em; border-color: #eaecf0; vertical-align: top; white-space: pre-wrap;"><br/></td></tr> <tr><td class="diff-marker"></td><td style="background-color: #f8f9fa; color: #202122; font-size: 88%; border-style: solid; border-width: 1px 1px 1px 4px; border-radius: 0.33em; border-color: #eaecf0; vertical-align: top; white-space: pre-wrap;"><br/></td><td class="diff-marker"></td><td style="background-color: #f8f9fa; color: #202122; font-size: 88%; border-style: solid; border-width: 1px 1px 1px 4px; border-radius: 0.33em; border-color: #eaecf0; vertical-align: top; white-space: pre-wrap;"><br/></td></tr> <tr><td class="diff-marker" data-marker="−"></td><td style="color: #202122; font-size: 88%; border-style: solid; border-width: 1px 1px 1px 4px; border-radius: 0.33em; border-color: #ffe49c; vertical-align: top; white-space: pre-wrap;"><div><del style="font-weight: bold; text-decoration: none;">Several </del>references <del style="font-weight: bold; text-decoration: none;">below</del>:</div></td><td class="diff-marker" data-marker="+"></td><td style="color: #202122; font-size: 88%; border-style: solid; border-width: 1px 1px 1px 4px; border-radius: 0.33em; border-color: #a3d3ff; vertical-align: top; white-space: pre-wrap;"><div><ins style="font-weight: bold; text-decoration: none;">== Some </ins>references: <ins style="font-weight: bold; text-decoration: none;">==</ins></div></td></tr> <tr><td class="diff-marker"></td><td style="background-color: #f8f9fa; color: #202122; font-size: 88%; border-style: solid; border-width: 1px 1px 1px 4px; border-radius: 0.33em; border-color: #eaecf0; vertical-align: top; white-space: pre-wrap;"><br/></td><td class="diff-marker"></td><td style="background-color: #f8f9fa; color: #202122; font-size: 88%; border-style: solid; border-width: 1px 1px 1px 4px; border-radius: 0.33em; border-color: #eaecf0; vertical-align: top; white-space: pre-wrap;"><br/></td></tr> <tr><td class="diff-marker"></td><td style="background-color: #f8f9fa; color: #202122; font-size: 88%; border-style: solid; border-width: 1px 1px 1px 4px; border-radius: 0.33em; border-color: #eaecf0; vertical-align: top; white-space: pre-wrap;"><div>Schultz, S. C., Shields, G. C. &amp; Steitz, T. A. (1990).</div></td><td class="diff-marker"></td><td style="background-color: #f8f9fa; color: #202122; font-size: 88%; border-style: solid; border-width: 1px 1px 1px 4px; border-radius: 0.33em; border-color: #eaecf0; vertical-align: top; white-space: pre-wrap;"><div>Schultz, S. C., Shields, G. C. &amp; Steitz, T. A. (1990).</div></td></tr> </table> Kay https://wiki.uni-konstanz.de/ccp4/index.php?title=Crystal_growth:_Protein-DNA_complexes&diff=1489&oldid=prev Cwilliam: New page: Tips suggested by the CCP4 community as the best way to crystallize a protein/DNA complex 1. One of the biggest variables is to screen different lengths of DNA 2. Good to try both blunt... 2009-07-20T11:51:43Z <p>New page: Tips suggested by the CCP4 community as the best way to crystallize a protein/DNA complex 1. One of the biggest variables is to screen different lengths of DNA 2. Good to try both blunt...</p> <p><b>New page</b></p><div>Tips suggested by the CCP4 community as the best way to crystallize a protein/DNA complex <br /> <br /> 1. One of the biggest variables is to screen different lengths of DNA<br /> <br /> 2. Good to try both blunt and sticky ends. Perhaps as a first attempt try sticky ends with complementary ends<br /> <br /> 3. Could try purification by gel filtration if the complex is tight. Probably easier, quicker, cheaper and just as successful just to mix and pray!<br /> <br /> 4. Screen for crystals using a wide variety of crystal screens<br /> Natrix, Nucleix, PEGS, Index, JCSG core, MPD screen, PEG ion screen, home made PEG salt screen (+/- Mg++)<br /> <br /> 5. DNA can be ordered from a wide range of suppliers. Could try using un-purified DNA for screening, purify your own or order HPLC purified DNA<br /> <br /> 6. Good to try a wide range of ratio’s for protein/DNA. Most popular seems to be a ratio of 1/1.2<br /> <br /> 7. Other useful tips?<br /> <br /> Protein DNA crystals can be very fragile so good idea to screen with cryoprotectant in the screen to save manipulations<br /> Test crystals with a gel to see if DNA is present<br /> Use of SAXS to get molecular envelop of complex<br /> <br /> <br /> Several references below:<br /> <br /> Schultz, S. C., Shields, G. C. &amp; Steitz, T. A. (1990).<br /> Crystallization of Escherichia coli catabolite gene activator protein with its DNA binding site. J. Mol.Biol. 213, 159–166.<br /> <br /> Tan et al, Crystallization of the Yeast MATa2/MCM1/DNA Ternary Complex: General Methods and Principles for Protein/DNA Cocrystallization J. Mol. Biol. (2000) 297, 947-959<br /> <br /> Cannone et al, Crystallization of bFGF-DNA aptamer complexes using a<br /> Sparse Matrix designed for protein–nucleic acid complexes Journal of Crystal Growth, 2001 232 (2001) 409–417<br /> <br /> Book chapter 8 from Crystallization of nucleic acids and proteins: a practical approach.<br /> Edited by A.Ducruix and R. Giege (Oxford University Press)</div> Cwilliam