Re: [ccp4bb] Off-topic: Native gel electrophoresis of basic proteins

Hi Ming, You can find a protocol for low pH native gel electrophoresis (as well for high pH and blue native) in the following book: Gel Electrophoresis of Proteins: A Practical Approach (Practical Approach Series) by B. D. Hames. http://www.amazon.com/Gel-Electrophoresis-Proteins-Practical-A

Re: [ccp4bb] Off-topic: Native gel electrophoresis of basic proteins

Ming, On that note, here is a Blue Native PAGE protocol that reportedly improves the separation of basic proteins. A friend and I have used it for native protein separation, but not necessarily for basic proteins. Word of warning: don't use histidine salts or Tris-HCl in the cathode buffer.

[ccp4bb] Summary: Dry shippers

Dear All, Thanks for the response. For those of you interested, the extra cost associated with shipping 'wet' is about 35 USD within the US, and about 50 to 100 dollar internationally. Extra paperwork and labels are needed when shipping wet. One of our users recommends shipping wet, especially i

Re: [ccp4bb] Handling protein crystals from 20% isopropanol

Hi! Vacuum oil can effectively stop evaporation completely if immediately dispensed onto the drop after the cover slip is removed from the well. 2x-5x the drop volume dispensed directly onto the drop creates a "drop within a drop", where air is prevented from reaching the inner drop, but the outer

Re: [ccp4bb] Off-topic: Native gel electrophoresis of basic proteins

You could try Coomassie Blue Native gel. It's a very neat technique and it worked for me on a couple of occasions. In one unfortunate case, it resulted in dissociation of a heterotetrameric complex, though. Artem > Dear CCP4 community, > > Sorry for the off-topic subject, but I would really appre

[ccp4bb] Off-topic: Native gel electrophoresis of basic proteins

Dear CCP4 community, Sorry for the off-topic subject, but I would really appreciate some suggestions and/or protocols relating to native gel electrophoresis of basic proteins. I have used a general acidic PAGE protocol for my protein, which has a PI of 9.5. Briefly, the protein was loaded onto a n

Re: [ccp4bb] Handling protein crystals from 20% isopropanol

Hello, My crystals in fact grew out of 20% peg4K and 20% isopropanol at 20 degrees and so were already effectively cryoprotected in the drop. Opening up the well at room temperature led to the crystals whirling around in the drop and made it hard to pick up a single crystal in the loop. The effect

Re: [ccp4bb] Dry shippers

I always used to just fill the shipper w/ LN2, and declare it Dangerous Goods. Never any trouble, save one time in Birmingham (AL), the FedEx person at the office intentionally tipped shipper 90 degrees, even though it was marked DG, Keep Upright, etc. !! Naturally, they got scared by all the "stea

[ccp4bb] Handling protein crystals from 20% isopropanol

Dear All: I have obtained good crystals from 20% isopropanol but this is my first time dealing with this agent. Help in handling the crystals, dealing with evaporation etc. is highly appreciated. Thank you Subbu

Re: [ccp4bb] Dry shippers

I'm interested to hear responses on this as well. We have shipped back and forth crystals a 100 times and never had a problem until our last trip to the synchrotron. The person at fed ex knocked our shipper onto it's side in order to verify the measurements. The fact that they did it with a big

Re: [ccp4bb] TLS refinement

Hi Artem, Thanks for your response, but how do you mean to do the ³normalization² with the other TLS related parameters. I have looked at parameters for each of my TLS groups (3 per monomer) dealing with translation and libration along 3 orthogonal axes and a correlation of the 2- a screw motion a

[ccp4bb] Proteopedia - because life is not 2D

Proteopedia - because life is not 2D Proteopedia is a new interactive web resource that presents 3D Biomolecular structures in an intuitive way by linking descriptive text to 3D images (see e.g., "HIV-1 protease" ). M

Re: [ccp4bb] Wilson plot from truncated.mtz

This has been said already but just to emphasise my view. 1) Measured intensities should never be "lost" and the TRUNCATED output should/ (and does by default) keep them. 2) TRUNCATE extracts Fs from Imeass in bins; it is far from perfect but we need amplitude estimates for some purposes and

[ccp4bb] Sharp/Resolve/ DM

This clears up some terrible confusions in my mind.. could one of you experts contribute a summary to the CCP4 Wiki or the crystallographic information site: http://strucbio.biologie.uni-konstanz.de/ccp4wiki/ Eleanor