Re: [ccp4bb] question regarding sequence numbering

Hi Tony, When I've had similar issues, I've numbered them sequentially (i.e. 7,8,9) and remarked in the PDB header that they are vector-derived sequence. I believe that is what the PDB ask you to do in situations like this (maybe they can comment?). If they are not numbered sequentially,

[ccp4bb] His-6 versus His-10 tag

Dear BB, We are planning the production of a protein for crystallization. From literature, we know that the construct with a 6-histidine tag crystallizes. However, for other biophysical measurements, we would prefer to have a 10-histidine tag. Does anyone has experience with His-6 versus

[ccp4bb] Kendrew Symposium, 16-17 Nov 2017, EMBL Heidelberg, Germany - Registration deadline October 5, 2017

Dear Colleagues, The 2-day Kendrew Symposium: "Revolutions in Structural Biology: Celebrating the 100th Anniversary of Sir John Kendrew" will take place on 16-17 November 2017 at EMBL Heidelberg. The aim of this event is to celebrate Sir John Kendrew 100th anniversary with outstanding

[ccp4bb] question regarding sequence numbering

Hi all, Here's a problem I haven't come across before. I'm working on a structure whose expression plasmid was designed to remove the first 9 amino acids from the protein of interest and to which an N-terminal tag was added. After cleaving the tag I am left with 3 amino acids (GPM) followed by

Re: [ccp4bb] His-6 versus His-10 tag

Hi Herman, I haven't done His-6 versus His-10 for the same protein, but have done that for different ones with success. However, if in His-6 containing protein structure the packing or folding is such that you don't see His-6 then it shouldn't matter it is 6 or 10. Just an opinion. Regards,

Re: [ccp4bb] His-6 versus His-10 tag

Hi Herman, I worked on a kinase, where moved from 6-His in the literature to 8-His, and it didn't impact crystallization or diffraction (which around 3Angs). Good Luck Partha On Tue, Sep 19, 2017 at 9:43 AM Oganesyan, Vaheh wrote: > Hi Herman, > > > > I haven’t done

Re: [ccp4bb] doubt regarding MR search model

Hello, Thanks everyone for your explanations. I have pasted the pointless output to provide more information. Best Solution: space group P 21 21 21 Laue group probability: 0.959 Systematic absence probability: 0.818 Total probability:

Re: [ccp4bb] His-6 versus His-10 tag

Hi Herman the survey article by Carson in Acta D DOI: 10.1107/S0907444906052024 from a decade ago may provide relevant information/insights. best wishes Savvas > On 19 Sep 2017, at 12:10, herman.schreu...@sanofi.com wrote: > > Dear BB, > > We

[ccp4bb] AW: [ccp4bb] question regarding sequence numbering

Hi Dave and Tony, Upon submission, the pdb checks the sequence and automatically generates comments about sequences derived from the expression vector. So you do not have to do anything. Given the issues many programs have with non-sequentially numbered residues, I would also number them

Re: [ccp4bb] His-6 versus His-10 tag

Dear Herman, I have crystallised the same protein with 6xHis and 12xHis and they both diffracted to high resolution. However, I have not seen these tags in the structure. Best wishes, Mohinder Sent from my iPhone > On 19 Sep 2017, at 12:10, herman.schreu...@sanofi.com wrote: > > Dear BB,

Re: [ccp4bb] doubt regarding MR search model

With Rs of 43/48% I don't think you can be sure that your spacegroup is right. You should always try all the spacegroup possibilities until you get a solution you are sure is right, i.e. that refines to Rs of around 35% or preferably even lower. More so in the case of screw axes, so try P222,

Re: [ccp4bb] His-6 versus His-10 tag

Option 1: make two constructs and compare :) Option 2: put the his10 on the n-term of the protein with a cleavage site In my experience it matters. I had cases where his4 crystallized and his6 did not. I also had the opposite experience. Artem On Sep 19, 2017 6:11 AM,

[ccp4bb] AW: [ccp4bb] doubt regarding MR search model

Dear Satvik, You only know if the space group is correct AFTER you solved your structure. With an Rwork/Rfree of 0.43/0.48, the space group is likely not correct. The way to solve this is to run MR in all possible space groups. Most, if not all MR programs have an option to do this

[ccp4bb] AW: [ccp4bb] His-6 versus His-10 tag

Dear everybody who reacted, It seems that when the His6 construct crystallizes, there is a fair chance that the his10 construct will crystallize as well. However, as usual in crystallography there is no guarantee, so I have added a TEV site to the construct in order to be able to remove the

Re: [ccp4bb] doubt regarding MR search model

Well - you haven't said what the sequence identity between model and your protein is, nor if you have a non-crystallographic translation. With low homology that R factor drop is acceptable and rebuilding can fix it, However if there is high homology you might expect better. But this sort of

[ccp4bb] Long-wavelength MX Beamline at Diamond (1st User Call)

The long-wavelength MX beamline I23 at Diamond Light Source has been added to the current Call for Proposals (deadline 1st Oct) for experiments from April 2018. This novel instrument can offer unique opportunities expanding the wavelength range for MX towards and beyond the sulphur K edge