Dear all, Thank you all for your valuable experiences, inputs and references!! I shall try them and hope for some good news! Its good to know there are so many examples of crystallization at such high concentrations.
A curious question - is it logical to say that if the protein is highly charged (either negative or positive), it is likely to be more soluble and resist crystallization due to electrostatic repulsion? Our protein has highly positively charged surface, although with some small negative patches. Following are some of the suggestions indicated: 1. Use water (will try) 2. Change buffers, pH temperature - (done) 3. Seeding (will try) 4. Methylating lysines (will try!!) 5. Surface entropy reduction (ongoing) 6. Optimize constructs (done) 7. Different ratios (done) 8. Keep concentrating (will try, the problem is yield is low!) 9. High salt concentrations (will try) 10. Organic solvents (though about it) 11. Mutations (ongoing) Thank you Regards Kavya On 2024-02-05 15:57, kavyashreem wrote: > Dear All, > > Has anyone worked on a protein which is highly soluble even at 80mg/ml? > > We have one such candidate, which does not precipitate even at 80mg/ml > instead forms phase separated globules in crystallization plate, which > eventually hardens over a period of 1 to 1.5 months (which is florescent > under UV microscope.) > > We tried screening at different pH, but failed to get any hits. > > Since we got few conditions in which the phase separated globules solidified, > we focused on them and expanded with 120mg/ml protein, still there were not > visible precipitates except for the phase separation. This has been a > challenging target so far. We have tried with different constructs, which > unfortunately are not soluble! > > Does POMs help in such cases? Or do you have any other suggestion. > > Thank you > > Regards > > Kavya > > CONFIDENTIAL: THIS EMAIL AND ANY FILES TRANSMITTED WITH IT ARE CONFIDENTIAL > AND INTENDED SOLELY FOR THE USE OF THE INDIVIDUAL OR GROUP TO WHOM THEY ARE > ADDRESSED. IF YOU HAVE RECEIVED THIS EMAIL IN ERROR, PLEASE NOTIFY THE SENDER > IMMEDIATELY AND DELETE THIS E-MAIL FROM YOUR SYSTEM. IT IS NOT AUTHORISED TO > COPY, FORWARD, OR IN ANY WAY REVEAL THE CONTENTS OF THIS MESSAGE TO ANYONE. > > CONFIDENTIAL: THIS EMAIL AND ANY FILES TRANSMITTED WITH IT ARE CONFIDENTIAL > AND INTENDED SOLELY FOR THE USE OF THE INDIVIDUAL OR GROUP TO WHOM THEY ARE > ADDRESSED. IF YOU HAVE RECEIVED THIS EMAIL IN ERROR, PLEASE NOTIFY THE SENDER > IMMEDIATELY AND DELETE THIS E-MAIL FROM YOUR SYSTEM. IT IS NOT AUTHORISED TO > COPY, FORWARD, OR IN ANY WAY REVEAL THE CONTENTS OF THIS MESSAGE TO ANYONE. > > ------------------------- > > 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 IS AN EXTERNAL MAIL AND **DID NOT** ORIGINATE FROM > **BLISC(NCBS/INSTEM/CCAMP)** CAMPUS SERVERS. PLEASE VERIFY THE CONTENT > CAREFULLY. PLEASE DO NOT SUBMIT USER CREDNTIALS ON ANY FORMS AND VERIFY THE > FROM ADDRESS AND URLs********* CONFIDENTIAL: This email and any files transmitted with it are confidential and intended solely for the use of the individual or group to whom they are addressed. If you have received this email in error, please notify the sender immediately and delete this e-mail from your system. It is NOT AUTHORISED to copy, forward, or in any way reveal the contents of this message to anyone. ######################################################################## 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/
