Hi Marco, In addition to what has already been suggested, you could also try co-crystallising with a peptide from the partner protein that was used for the existing structure. Specifically, use a peptide from and interacting surface, which might help with the crystal packing, as well as stabilising your protein (if this is an issue).
Cheers, Sara ________________________________ From: CCP4 bulletin board <CCP4BB@JISCMAIL.AC.UK> on behalf of Marco Bravo <0000d0eb7bee83ae-dmarc-requ...@jiscmail.ac.uk> Sent: 10 January 2025 01:00 To: CCP4BB@JISCMAIL.AC.UK <CCP4BB@JISCMAIL.AC.UK> Subject: [ccp4bb] Help crystalizing protein CAUTION: This email originated from outside of the ICR. Do not click links or open attachments unless you recognize the sender's email address and know the content is safe. Hello all, I am seeking some advice or ideas to crystalize a nuclease protein. It exists within a heterodimer, homodimer, and maybe monomer as well depending on the species from which the protein is in. The protein has been shown to have nuclease activity on its own depending on the species. I am screening the protein from three different species and have setup so many crystal high-throughput crystal screens with and without DNA. I never get protein crystals for these proteins, I have done thermal shift assays to identify a buffer that stabilizes the protein better than the storage buffer and found some conditions that increased the melting temp by 4-5 degrees. I screened the protein again in these new stabilizing buffers but still have not gotten any hits. There are protein structures for the protein in the pdb with its partner protein in the heterodimer formation. I have experience screening other proteins and getting several hits per kit so I know what to expect for a successful screen. Any advice would be helpful, I want to retain the full-length protein for structural studies. I would appreciate any ideas or advice for moving forward and trying to obtain crystals. Thanks! Marco ######################################################################## To unsubscribe from the CCP4BB list, click the following link: https://eur01.safelinks.protection.outlook.com/?url=https%3A%2F%2Fwww.jiscmail.ac.uk%2Fcgi-bin%2FWA-JISC.exe%3FSUBED1%3DCCP4BB%26A%3D1&data=05%7C02%7Csara.silva%40ICR.AC.UK%7C58819c67339344fa9fd208dd311251ec%7C6138f7b9eeea47f7a06bb90a692f238e%7C0%7C0%7C638720676940116701%7CUnknown%7CTWFpbGZsb3d8eyJFbXB0eU1hcGkiOnRydWUsIlYiOiIwLjAuMDAwMCIsIlAiOiJXaW4zMiIsIkFOIjoiTWFpbCIsIldUIjoyfQ%3D%3D%7C0%7C%7C%7C&sdata=qw61YuVTLTdPOwR28t92rBl9N8tPGyQO8bkDk85mwrs%3D&reserved=0<https://www.jiscmail.ac.uk/cgi-bin/WA-JISC.exe?SUBED1=CCP4BB&A=1> This message was issued to members of https://eur01.safelinks.protection.outlook.com/?url=http%3A%2F%2Fwww.jiscmail.ac.uk%2FCCP4BB&data=05%7C02%7Csara.silva%40ICR.AC.UK%7C58819c67339344fa9fd208dd311251ec%7C6138f7b9eeea47f7a06bb90a692f238e%7C0%7C0%7C638720676940138361%7CUnknown%7CTWFpbGZsb3d8eyJFbXB0eU1hcGkiOnRydWUsIlYiOiIwLjAuMDAwMCIsIlAiOiJXaW4zMiIsIkFOIjoiTWFpbCIsIldUIjoyfQ%3D%3D%7C0%7C%7C%7C&sdata=Bkt0M8bTtnpPKK0pb%2FH6etGE7%2FzOn0ybb6i6jGRzy1Q%3D&reserved=0<http://www.jiscmail.ac.uk/CCP4BB>, a mailing list hosted by https://eur01.safelinks.protection.outlook.com/?url=http%3A%2F%2Fwww.jiscmail.ac.uk%2F&data=05%7C02%7Csara.silva%40ICR.AC.UK%7C58819c67339344fa9fd208dd311251ec%7C6138f7b9eeea47f7a06bb90a692f238e%7C0%7C0%7C638720676940151169%7CUnknown%7CTWFpbGZsb3d8eyJFbXB0eU1hcGkiOnRydWUsIlYiOiIwLjAuMDAwMCIsIlAiOiJXaW4zMiIsIkFOIjoiTWFpbCIsIldUIjoyfQ%3D%3D%7C0%7C%7C%7C&sdata=ICgXh68ayN%2BzRJHBAXhEmJJmrrwoUN1p62ePmyzarCk%3D&reserved=0<http://www.jiscmail.ac.uk/>, terms & conditions are available at https://eur01.safelinks.protection.outlook.com/?url=https%3A%2F%2Fwww.jiscmail.ac.uk%2Fpolicyandsecurity%2F&data=05%7C02%7Csara.silva%40ICR.AC.UK%7C58819c67339344fa9fd208dd311251ec%7C6138f7b9eeea47f7a06bb90a692f238e%7C0%7C0%7C638720676940162704%7CUnknown%7CTWFpbGZsb3d8eyJFbXB0eU1hcGkiOnRydWUsIlYiOiIwLjAuMDAwMCIsIlAiOiJXaW4zMiIsIkFOIjoiTWFpbCIsIldUIjoyfQ%3D%3D%7C0%7C%7C%7C&sdata=htuvIVbxjgB%2F5ksGDMX0wp7%2BAtoNY2qcC5hATYEuDr8%3D&reserved=0<https://www.jiscmail.ac.uk/policyandsecurity/> The Institute of Cancer Research: Royal Cancer Hospital, a charitable Company Limited by Guarantee, Registered in England under Company No. 534147 with its Registered Office at 123 Old Brompton Road, London SW7 3RP. This e-mail message is confidential and for use by the addressee only. If the message is received by anyone other than the addressee, please return the message to the sender by replying to it and then delete the message from your computer and network. ######################################################################## To unsubscribe from the CCP4BB list, click the following link: https://www.jiscmail.ac.uk/cgi-bin/WA-JISC.exe?SUBED1=CCP4BB&A=1 This message was issued to members of www.jiscmail.ac.uk/CCP4BB, a mailing list hosted by www.jiscmail.ac.uk, terms & conditions are available at https://www.jiscmail.ac.uk/policyandsecurity/
