[ccp4bb] Isolation of protein-protein complexes.

2014-01-21 Thread David Briggs
Dear all, sorry for the slightly off topic post, I have 2 proteins that have been shown to interact, by multiple groups, and by multiple techniques - namely ELISA, SPR and DPI. The Kd of the interaction as determined by SPR is on the order of 1 nM. I would very much like to crystallise this

Re: [ccp4bb] Isolation of protein-protein complexes.

2014-01-21 Thread Philippe Leone
Dear Dave, Indeed, an interaction in the nM range is strong, but its dynamic is important. If the SPR sensorgram, if published, looks like a 'crenel', this indicates high association/dissociation rates. In that case, your complex can dissociate during the GF run. You can try crystallisation

Re: [ccp4bb] Isolation of protein-protein complexes.

2014-01-21 Thread Bosch, Juergen
Hey David, the 1 nM Kd by SPR sounds fishy to me - did you do these measurements yourself ? Unless this is an antibody-protein complex or nanobody-protein complex or protein-small molecule complex, the value is too low (for a normal PPI, I would expect 50 -500 nM). Possible explanation for the

Re: [ccp4bb] Isolation of protein-protein complexes.

2014-01-21 Thread Keller, Jacob
Subject: [ccp4bb] Isolation of protein-protein complexes. Dear all, sorry for the slightly off topic post, I have 2 proteins that have been shown to interact, by multiple groups, and by multiple techniques - namely ELISA, SPR and DPI. The Kd of the interaction as determined by SPR is on the order

Re: [ccp4bb] Isolation of protein-protein complexes.

2014-01-21 Thread David Briggs
Hi all, Thanks for the incredibly fast and helpful responses, both on and off list. To provide some further information. I have tried the SEC with several buffers, including buffers identical to the SPR and DPI experiments. To-date all SEC and SPR experiments have been conducted at room

Re: [ccp4bb] Isolation of protein-protein complexes.

2014-01-21 Thread Eugene Osipov
@JISCMAIL.AC.UK *Subject:* [ccp4bb] Isolation of protein-protein complexes. Dear all, sorry for the slightly off topic post, I have 2 proteins that have been shown to interact, by multiple groups, and by multiple techniques - namely ELISA, SPR and DPI. The Kd of the interaction as determined

Re: [ccp4bb] Isolation of protein-protein complexes.

2014-01-21 Thread Enrico Stura
that was in the buffer used for the other binding studies, but not in your SEC buffer? JPK *From:* CCP4 bulletin board [mailto:CCP4BB@JISCMAIL.AC.UK] *On Behalf Of *David Briggs *Sent:* Tuesday, January 21, 2014 10:52 AM *To:* CCP4BB@JISCMAIL.AC.UK *Subject:* [ccp4bb] Isolation of protein