Re: [ccp4bb] intermolecular dissulphides
Dear Eleanor, sorry for the late reply. I found a similar case (id: 2OEF, Cys 92). I mutated the Cys to Ser, but it did not change anything (xtal conditions and quality remained the same). In solution the protein is a monomer. However, adding 1 mM DTT resulted in a significant increase in crystal quality (larger 3D crystals with better resolution). I don't know why, though... Cheers, Johannes 2017-02-01 20:45 GMT+01:00 Tom Peat : > Hello Eleanor, > > > We found some intermolecular vicinal disulfides recently that we think are > 'real'. This class of proteins forms tetramers and in one version we find > these intermolecular disulfides across molecules. We did some tests and > found that oxidation or reduction has an effect on the stability of the > protein. We also saw this was consistent across multiple space groups. If > you would like to have a look, they were just released: 5HY0, 5HY2, 5HY4. > > As a comparison to another protein in this fold class that doesn't have > the disulfide is 5HWE. > > > cheers, tom > > > Tom Peat > Proteins Group > Biomedical Program, CSIRO > 343 Royal Parade > Parkville, VIC, 3052 > +613 9662 7304 <+61%203%209662%207304> > +614 57 539 419 > tom.p...@csiro.au > -- > *From:* CCP4 bulletin board on behalf of Eleanor > Dodson > *Sent:* Thursday, February 2, 2017 2:17 AM > *To:* CCP4BB@JISCMAIL.AC.UK > *Subject:* [ccp4bb] intermolecular dissulphides > > Does anyone know of examples of these? > I have found one - 2WQW with these SSBOND records > 2WQW > SSBOND 1 CYS A 206CYS A 227 1555 6556 > 2.07 > SSBOND 2 CYS B 206CYS B 227 1555 5556 > 2.15 > > We seem to have one but it would have to form after crystalisation? > > Eleanor > > >
Re: [ccp4bb] intermolecular dissulphides
Hello Eleanor, We found some intermolecular vicinal disulfides recently that we think are 'real'. This class of proteins forms tetramers and in one version we find these intermolecular disulfides across molecules. We did some tests and found that oxidation or reduction has an effect on the stability of the protein. ?We also saw this was consistent across multiple space groups. If you would like to have a look, they were just released: 5HY0, 5HY2, 5HY4. As a comparison to another protein in this fold class that doesn't have the disulfide is 5HWE. cheers, tom Tom Peat Proteins Group Biomedical Program, CSIRO 343 Royal Parade Parkville, VIC, 3052 +613 9662 7304 +614 57 539 419 tom.p...@csiro.au From: CCP4 bulletin board on behalf of Eleanor Dodson Sent: Thursday, February 2, 2017 2:17 AM To: CCP4BB@JISCMAIL.AC.UK Subject: [ccp4bb] intermolecular dissulphides Does anyone know of examples of these? I have found one - 2WQW with these SSBOND records 2WQW SSBOND 1 CYS A 206CYS A 227 1555 6556 2.07 SSBOND 2 CYS B 206CYS B 227 1555 5556 2.15 We seem to have one but it would have to form after crystalisation? Eleanor
Re: [ccp4bb] intermolecular dissulphides
Does this count as an example? grep SSBOND /a/pdb/pdb4e9m.ent SSBOND 1 CYS A 39CYS B 39 1555 1555 2.05 SSBOND 2 CYS C 39CYS D 39 1555 1555 2.04 SSBOND 3 CYS E 39CYS F 39 1555 1555 2.03 The A.U. contains three domain-swapped dimers. The cys are not on the swapped helix but the swapping fortuitously brings the same cys in the two molecules into proximity to make a disulfide. There are two or three other x-ray structures that show the same domain-swapped, disulfide-clinched dimer in different packing. However an NMR structure shows it to be monomeric in solution, based on estimated tumbling speed since nmr restraints might not distinguish inter- from intramolecular contacts in a domain-swapped dimer. The cys is not conserved, and although this protein is expected to oligomerize, the putative oligomerization domain is not included in this construct. On 02/01/2017 10:17 AM, Eleanor Dodson wrote: Does anyone know of examples of these? I have found one - 2WQW with these SSBOND records 2WQW SSBOND 1 CYS A 206CYS A 227 1555 6556 2.07 SSBOND 2 CYS B 206CYS B 227 1555 5556 2.15 We seem to have one but it would have to form after crystalisation? Eleanor
Re: [ccp4bb] intermolecular dissulphides
Thank you for all this information, and especially Paul for having a good search query - I had got nothing useful from any I tried at PDBe or RSCB. No idea why it has been formed - the fold is identical to a high resolution example (1.3A) where the disulphide links the N & C termini within a single molecule. Except for 2-3 residues which cause the domain swap the CA rmsd is < 2. It must be a crystallisation artefact I think,. or my mistake in interpretation... This new data is lower resolution but after omitting the hings region the density is pretty reliable.. Eleanor On 1 February 2017 at 16:13, Eleanor Dodson wrote: > Thank you VERY much - how did you generate that? > E > > On 1 February 2017 at 16:10, Paul Emsley > wrote: > >> On 01/02/2017 15:17, Eleanor Dodson wrote: >> >>> Does anyone know of examples of these? >>> >> >> Here's a rough list - not all of them are real. >> >> P. >> >> >> >> >
[ccp4bb] AW: [ccp4bb] intermolecular dissulphides
Dear Eleanor, I did not check the pdb file you mentioned, but I have had a case like that. The protein formed a complex of 8 large and 8 small subunits with internal 422 symmetry. There was a disulfide link across the internal twofolds and in one of the crystal forms we got, this internal twofold came on top of a crystallographic twofold. At the time, I did not know about these sophisticated SSBOND records (if they existed at the time), so I assigned a very small van der Waals radius to the SGs to solve the repulsion problem. So, if you have a dimer with an existing disulfide link across the twofold axis, this twofold axis may become a crystallographic twofold and there is no need for the disulfide bond to form afterwards. Best regards, Herman Von: CCP4 bulletin board [mailto:CCP4BB@JISCMAIL.AC.UK] Im Auftrag von Eleanor Dodson Gesendet: Mittwoch, 1. Februar 2017 16:18 An: CCP4BB@JISCMAIL.AC.UK Betreff: [ccp4bb] intermolecular dissulphides Does anyone know of examples of these? I have found one - 2WQW with these SSBOND records 2WQW SSBOND 1 CYS A 206CYS A 227 1555 6556 2.07 SSBOND 2 CYS B 206CYS B 227 1555 5556 2.15 We seem to have one but it would have to form after crystalisation? Eleanor
Re: [ccp4bb] intermolecular dissulphides
I solved a structure years ago (1L31 and a couple related entries) which had intermolecular oligomerizing disulfides in some of the crystals (reciprocal C23-C27’ and C23’-C27). I thought at the time that they seemed too “deliberate” to be just an artifact, but others thought that since the enzyme was intracellular, the disulfides would not be physiologically relevant. I still think they’re too perfect to be artefactual, and since the protein comes from Methanobacterium thermoautotrophicum, all usual bets might be off. I’ve always wondered about it, although I am not sure how profound the consequences are. I would love to hear peoples’ two cents on this. Jacob From: CCP4 bulletin board [mailto:CCP4BB@JISCMAIL.AC.UK] On Behalf Of Eleanor Dodson Sent: Wednesday, February 01, 2017 10:18 AM To: CCP4BB@JISCMAIL.AC.UK Subject: [ccp4bb] intermolecular dissulphides Does anyone know of examples of these? I have found one - 2WQW with these SSBOND records 2WQW SSBOND 1 CYS A 206CYS A 227 1555 6556 2.07 SSBOND 2 CYS B 206CYS B 227 1555 5556 2.15 We seem to have one but it would have to form after crystalisation? Eleanor
[ccp4bb] intermolecular dissulphides
Does anyone know of examples of these? I have found one - 2WQW with these SSBOND records 2WQW SSBOND 1 CYS A 206CYS A 227 1555 6556 2.07 SSBOND 2 CYS B 206CYS B 227 1555 5556 2.15 We seem to have one but it would have to form after crystalisation? Eleanor
