Re: [ccp4bb] intermolecular dissulphides

[Johannes Cramer]

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

[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
+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

[Edward A. Berry]

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

[Eleanor Dodson]

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.
>>
>>
>>
>>
>

Re: [ccp4bb] intermolecular dissulphides

[Keller, Jacob]

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