[ccp4bb] [off topic] Recovering pET expression plasmid from BL21 strain

Dear all, I'm sure this issue comes up very often, but for the first time in our lab we need to recover a pET-type expression plasmid from a BL21-like E. coli strain (NEB's T7 Express). I know a RecA+ strain is not suitable for plasmid production, but the basic plan is to grow and mini-prep the

Re: [ccp4bb] Validation of structure prediction

Hello, I think it is perfectly "valid" to run AlphaFold2 / RosettaFold / etc. models through geometry validation servers (aside from reporting their intrinsic quality indicators like TM-score and pLDDT distribution, as mentioned above by Randy Read). Since most experimental structures have a small

[ccp4bb] [just for fun] please vote #JBCMethodsMadness #TeamXRC

just one hour left !! https://twitter.com/jbiolchem/status/1374030429729271811?s=20 -- Dr. Javier M. González Instituto de Bionanotecnología del NOA (INBIONATEC-CONICET) Universidad Nacional de Santiago del Estero (UNSE) RN9, Km 1125. Villa El Zanjón. (G4206XCP) Santiago del Estero. Argentina

Re: [ccp4bb] To solve the problem of an extremely asymmetric peak shape obtained from gel filtration chromatography

Hello, I agree with Roger, you should definitely try to increase the salt concentration to get rid of non specific binding impurities. And if that doesn't work, you can try purifying your protein under denaturing conditions by adding one refolding step in the column. Good luck, Javier On Wed, Dec

Re: [ccp4bb] Macports and Fink - failed building open source pymol on MacOS Catalina

; > On Sat, Jun 13, 2020 at 4:45 PM Javier Gonzalez wrote: > >> Hello, >> I'm attempting to build pymol on a laptop running MacOS Catalina 10.15.5, >> Processor 2.8GHz Quad-Core Intel i7, Graphics Intel Iris Pro 1536 MB >> I downloaded the latest version of Xcode 11.1

[ccp4bb] Macports and Fink - failed building open source pymol on MacOS Catalina

Hello, I'm attempting to build pymol on a laptop running MacOS Catalina 10.15.5, Processor 2.8GHz Quad-Core Intel i7, Graphics Intel Iris Pro 1536 MB I downloaded the latest version of Xcode 11.1 I tried from Fink (fink install pymol-py27) and Macports (sudo port install pymol), as indicated

Re: [ccp4bb] Vote for cryoEM

1. We need renowned scientists on Twitter because it is one of the best ways to spread words worth reading, mostly at these times when so many pseudo-scientists are influencers on social media (or worse, some even have a show on Netflix). 2. I'm voting for Mass Spec right now Cheers, @_biojmg On

Re: [ccp4bb] Protein cavity volume calculation

Hi Seema, You can also have a look at https://www.caver.cz/ they´ve done a great job with the CAVER program to calculate, analyze and visualize channels and tunnels in macromolecules. Best wishes, Javier On Sat, Jun 15, 2019 at 7:57 PM Seema H Irani wrote: > > I was wondering if anyone could

Re: [ccp4bb] Re-using 96-well crystallization plates

Hello Nemanja, I used to wash and reuse glass plates for neutron crystallography. Of course glass is sturdier than polystyrene, but I can't think of any protein stain that would resist a treatment with detergent, then a strong base (say 0.1M NaOH) and finally a strong acid (say 0.1M HCl)...

Re: [ccp4bb] Off topic question

Hi Reza, happy new year! The choice would depend on your alignment (aminoacid or nucleotides? are the sequences closely or distantly related? is it a large alignment? are there many gaps?)... Anyway, I think the safest, unbiased way to determine a group of outliers might be to compute a

Re: [ccp4bb] nonenzymatic removal of His tag?

Another option would be to use a protease immobilized in some resin (for instance GST-tagged TEV protease bound to GSH-Agarose resin, in column or in batch), so that no protease would be free to co-purify with your protein or peptide. HTH, Javier On Thu, Sep 20, 2018 at 5:35 PM, Patrick Loll

Re: [ccp4bb] Zn+2 proteases

Hello Jiri, I wouldn't choose neither of them, but Co(II). NMR spectroscopists have been using Co(II) to probe Zn(II) sites in proteins for ages... Check out Bertini (*DOI: *10.1021/ar00092a002 ), or Vallee (*DOI: * 10.1021/bi00476a001

Re: [ccp4bb] capillary counterdiffusion agarose plug

Hello Georg, You can melt the agarose in a glass container using a microwave oven. Then connect your capillary tube to small syringe, using a plastic tubing or adapter. Use some parafilm if the tubing diameters do not fit exactly, there should be no leaks. Then slowly pull the plunger to take

Re: [ccp4bb] software/web server to determine ligand volume

You can calculate volumes and much more, here: http://www.molinspiration.com/cgi-bin/properties Javier On Wed, Aug 13, 2014 at 12:06 AM, sreetama das somon_...@yahoo.co.in wrote: Dear all, Is there any software or web server available to calculate the volume of a ligand if the ligand

Re: [ccp4bb] Google Gets it Right

Interestingly, that molecule has the opposite configuration. The actual modelhttp://upload.wikimedia.org/wikipedia/commons/6/6e/Molecular_model_of_Penicillin_by_Dorothy_Hodgkin_%289663803982%29.jpghas a posting saying (wrong absolute configuration). I wonder what's the story behind it... On

[ccp4bb] Usage of gels in protein crystallography

Dear CCP4BBers, I would very much appreciate any information or resources regarding usage of gels in order to achieve supersaturation/crystallization through liquid diffusion. It appears to me that, although this crystallization method is usually claimed as a powerful technique, it is very

[ccp4bb] Are there.cif dictionaries for deuterated aminoacids for Coot?

Dear all, I'm refining a neutron structure and I would like to know whether there are available geometry dictionaries for deuterated aminoacids, in order to input them into Coot. At least the version I have (Coot 0.7, revision 4459) is unable to handle aminoacids whose exchangeable H atoms are

Re: [ccp4bb] vitrification vs freezing

Hi Sebastiano, I think the term vitrified crystal could be understood as a very nice oxymoron (http://www.oxymoronlist.com/), but it is essentially self-contradictory and not technically correct. As Ethan said, vitrify means turn into glass. Now, a glass state is a disordered solid state by

[ccp4bb] Job Posting - Laboratory Specialist in NMR, São Paulo University, Brazil

This is a forwarded message. For inquires please contact chsfa...@iq.usp.bror robe...@iq.usp.br ___ The Central Analítica at the Instituto de Química, Universidade de São Paulo has an opening for a Laboratory Specialist in NMR. The position is associated with the acquisition of an 800 MHz

Re: [ccp4bb] how to remove part of data with bad signal to noise ratio

I would also make sure that the spacegroup is correct. If you have instead P222, P2212, etc, you might find the solution at low resolution but the problem would become evident during advanced refinement steps, such as a stuck high Rfree or a noisy difference map. Good luck, Javier. On Sat, May

Re: [ccp4bb] LIGPLOT or similar

Mark, you might want to try PDBsum (http://www.ebi.ac.uk/pdbsum/), it will generate a LIGPLOT output for a given PDB code entered, which can be downloaded as high resolution .pdf or .ps Best, Javier On Wed, Aug 25, 2010 at 2:49 PM, Tim Gruene t...@shelx.uni-ac.gwdg.de wrote: Hello Mark, if

Re: [ccp4bb] attachments

Dear all, I've just created a group on Facebook named CCP4 Stuff. This would be a good place to upload as much random stuff as you want (images, videos, links, comments, etc) that nobody will care about after a couple of days. All you need is to join (this is not an issue, considering that most

Re: [ccp4bb] How could I Extract Iron from Protein?

Since you have only Asp and His ligands coordinating the Fe ion, dialysis against, say, 0.1 M Citrate at pH 5 will do the trick. Citrate will chelate the Fe(III) (if the protein has Fe(II) it will be oxidized during dialysis due to air oxygen) avoiding precipitation of Fe(OH)3 and the acid will