Re: [ccp4bb] Efforts been made to solve protein (soluble) aggregation

2014-03-06 Thread Bosch, Juergen
Co expression of binding partner perhaps ? And then tagging the other protein partner ? Jürgen .. Jürgen Bosch Johns Hopkins Bloomberg School of Public Health Department of Biochemistry Molecular Biology Johns Hopkins Malaria Research Institute 615 North Wolfe Street, W8708

Re: [ccp4bb] Peptide solubility issues

2014-03-05 Thread Bosch, Juergen
For that purpose we’ve developed this expression system that may or may not be useful to your specific problem: http://www.ncbi.nlm.nih.gov/pubmed/23996492 J Mol Recognit.http://www.ncbi.nlm.nih.gov/pubmed/23996492# 2013 Oct;26(10):496-500. doi: 10.1002/jmr.2292. Development of a multifunctional

Re: [ccp4bb] Sister CCPs

2014-02-13 Thread Bosch, Juergen
Let me pick up Eleanor’s comment: is there something like a crystallographer today ? I mean in the true sense ? I think as a “crystallographer” you won’t be able to survive the next decade, you need to diversify your toolset of techniques as pointed out in this article

Re: [ccp4bb] Sister CCPs

2014-02-13 Thread Bosch, Juergen
, please do not discourage your colleagues, particularly very junior ones, from posting to the CCP4BB. Some of the questions may appear quaint or irrelevant but it is easy to simply ignore topics that are of no interest! Eugene On 13 February 2014 14:41, Bosch, Juergen jubo

Re: [ccp4bb] How to find the unfound part of a big protein

2014-02-13 Thread Bosch, Juergen
Hi Bingfa, first suggestion would be to process your data with the anomalous flag turned on - just in case you happen to have some metal bound coincidentally and you happen to have collected at a decent wavelength to pickup some anomalous scattering. ~30% of proteins in the PDB have a metal

Re: [ccp4bb] off-topic: bug busting

2014-02-05 Thread Bosch, Juergen
If heat is a concern, you can place your Emulsiflex into a bin with lots of ice. We have the version with cooling after lysis, but if you are paranoid you can cool the whole thing. It has the footprint of a 15” MacBook Pro but it’s way more expensive than that :-) Jürgen

Re: [ccp4bb] identifying protein crystals via visible light only?

2014-02-04 Thread Bosch, Juergen
[Advertisement on] http://www.janscientific.com/#!/gallery [Advertisement off] Jürgen .. Jürgen Bosch Johns Hopkins University Bloomberg School of Public Health Department of Biochemistry Molecular Biology Johns Hopkins Malaria Research Institute 615 North Wolfe Street,

Re: [ccp4bb] off-topic: bug busting

2014-02-04 Thread Bosch, Juergen
Emulsiflex C5 alternative freeze-thawing, the old way we used to do this spiking with some Lysozyme or grinding it in a LN2 cooled mortar, is very effective and pretty cost effective, assuming you don’t pay for the LN2 Jürgen .. Jürgen Bosch Johns Hopkins University

Re: [ccp4bb] Isolation of protein-protein complexes.

2014-01-21 Thread Bosch, Juergen
Hey David, the 1 nM Kd by SPR sounds fishy to me - did you do these measurements yourself ? Unless this is an antibody-protein complex or nanobody-protein complex or protein-small molecule complex, the value is too low (for a normal PPI, I would expect 50 -500 nM). Possible explanation for the

Re: [ccp4bb] How to show the ligands (both as sticks and sphere) as shown here in Pymol

2014-01-19 Thread Bosch, Juergen
Hi Wei, make two objects of your ligand, display one as sphere and the other as stick. Then change the transparency setting for the spheres. Jürgen On Jan 19, 2014, at 11:40 PM, Wei Shi wei.shi...@gmail.commailto:wei.shi...@gmail.com wrote: Hi all, Please see attached Fig where they show the

Re: [ccp4bb] Protein concentration form chromatograms

2014-01-15 Thread Bosch, Juergen
Step1: visit Protparam tool and CP your sequence, scroll down until you find the extinction coefficient part. If OD280 is not close to 1 then make sure to take that into account in your chromatogram say it is 0.7 step2: look at your mAU's If your peak is 1000 mAU then you have 0.7 mg/ml in that

Re: [ccp4bb] Heavy atom sites

2013-12-16 Thread Bosch, Juergen
Didn't mlphare use to print those values in the log file ? Jürgen On Dec 15, 2013, at 4:29 PM, David Schuller dj...@cornell.edumailto:dj...@cornell.edu wrote: I have some SIRAS data of a known structure. I want to get the isomorphous and anomalous occupancy and phasing power from my data.

Re: [ccp4bb] A question on protein microheterogenity for crystalization

2013-12-14 Thread Bosch, Juergen
Hi Acoot, since they behave differently on IEX, they are different - you would introduce heterogeneity into your crystallization setup, which usually is not a good idea for crystallization in general. Are you using Benzonucleases by any chance in your preparation and if not, that may explain

Re: [ccp4bb] Wrong Space Group?

2013-12-13 Thread Bosch, Juergen
2.8 is not a terrible resolution to try out Buccaneer or Parrot. Your terrible R-factor might be due to a shift in residues perhaps ? Your After-building map does not show much of a side chain density to judge if you are in frame or off. But the elongated helix is in my eyes convincing enough.

Re: [ccp4bb] Off topic: Problem in reproducing crystal screen

2013-12-09 Thread Bosch, Juergen
another option might be the age of the screen perhaps ? very silly suggestion/question: your pH meter is calibrated before you make your measurements right ? If you have a nano-pH probe (anything that can be added to a 96well reservoir well) you could test what the final pH of your composition

Re: [ccp4bb] How can I find the other molecule in the asymmetric unit?

2013-11-21 Thread Bosch, Juergen
What is the solution to this? Hi Meisam, you have it, it is just three molecules in the asu. Look at the overall crystal lattice packing and see if you have contacts supporting each molecule. Generate a large representation of your symmetry mates, I suspect you have a channel in your crystal

Re: [ccp4bb] PDB structure validation

2013-10-31 Thread Bosch, Juergen
yes, indeed. visit this site and make sure you get all green lights http://molprobity.biochem.duke.edu Jürgen On Oct 31, 2013, at 11:25 AM, Debasish Chattopadhyay wrote: I was wondering if there is a way to generate a PDB validation report before depositing the coordinates so that one can go

Re: [ccp4bb] PDB structure validation

2013-10-31 Thread Bosch, Juergen
You do use Coot and look at the density plots right ? Phenix will essentially do the same but in text form (unless you use the GUI). You can also do this with Solve/Resolve and you can write your own script with CCP4 available tools to do the same RSR fit diagram. Jürgen On Oct 31, 2013, at

Re: [ccp4bb] Unrelated - Emulsiflex C3 leaking

2013-10-30 Thread Bosch, Juergen
We've had that leak as well some time ago. Dismantle and reassemble it is the trick. Just a slight angle when tightening it will lead to that problem. This is usually the case because of overtughtening the reservoir to the main part of the machine. Keep it always straight when closing it again.

Re: [ccp4bb] Biacore/SPR

2013-10-28 Thread Bosch, Juergen
Protein-protein interaction? protein-small molecule interactions ? Epitope mapping of mABs ? Could you specify what you would like to do, as different models are good for different things. Jürgen On Oct 28, 2013, at 10:08 AM, Gang Dong wrote: Dear all, Could anyone tell me your experience with

Re: [ccp4bb] Biacore/SPR

2013-10-28 Thread Bosch, Juergen
BiaCore 3000 or if you can afford the T200. Proteon XPR36 would be a cheaper Option and should work as well. Jürgen On Oct 28, 2013, at 10:54 AM, Gang Dong wrote: We mainly measure protein-protein interactions (sometimes protein-small molecules). Thanks! _Gang From: Bosch, Juergen [mailto:jubo

Re: [ccp4bb] a problem when using NOMACHINE Player from OSX 10.7.4

2013-09-18 Thread Bosch, Juergen
check out the hidden .nx folder in your home directory. You should have a folder named config. You may have a .nxs file or .cfg file there just edit the file with a text editor. You probably need to change this value: option key=Resolution value=1440x900 / If you have a retina MacBook I guess

Re: [ccp4bb] Adding ligand to crystallization drop

2013-09-12 Thread Bosch, Juergen
Have a look at these two papers: http://www.ncbi.nlm.nih.gov/pubmed/17004709 http://www.ncbi.nlm.nih.gov/pubmed/19929835 When you say high DMSO, how much is that in % ? Do you know if your crystals survive that much percentage DMSO even without ligand ? Jürgen .. Jürgen

Re: [ccp4bb] over-expression strategies

2013-09-12 Thread Bosch, Juergen
Is your gene of interest smaller than 750bp ? Then I would synthesize the gene with IDT and optimize it for E.coli that's 139$ Jürgen .. Jürgen Bosch Johns Hopkins Bloomberg School of Public Health Department of Biochemistry Molecular Biology Johns Hopkins Malaria Research

Re: [ccp4bb] Adding ligand to crystallization drop

2013-09-12 Thread Bosch, Juergen
But you have not tested in the beam if the visually happy crystals diffract right ? I would do this first before consuming your precious ligands in worthless drops. Jürgen On Sep 12, 2013, at 10:34 AM, Christopher Browning wrote: Right now I've tried 5, 10 and 16% DMSO (due to variations in

Re: [ccp4bb] Resolution, R factors and data quality

2013-08-28 Thread Bosch, Juergen
What a statement ! Give reviewers maps, I agree however, what if the reviewer has no clue of these things we call structures ? I think for those people table 1 might still provide some justification. I would argue it should go into the supplement at least. Jürgen Sent from my iPad On Aug

[ccp4bb] Quick resolution cutoff survey

2013-08-28 Thread Bosch, Juergen
Since we keep discussing resolution cutoffs and the benefits of not to include all data etc. I thought I would crowd source your opinion on this particular data set. processed with XDS, here's the XSCALE.LP output: SUBSET OF INTENSITY DATA WITH SIGNAL/NOISE = -3.0 AS FUNCTION OF RESOLUTION

Re: [ccp4bb] Quick resolution cutoff survey

2013-08-28 Thread Bosch, Juergen
Health Department of Biochemistry Molecular Biology Johns Hopkins Malaria Research Institute 615 North Wolfe Street, W8708 Baltimore, MD 21205 Office: +1-410-614-4742 Lab: +1-410-614-4894 Fax: +1-410-955-2926 http://lupo.jhsph.edu On Aug 28, 2013, at 11:32 AM, Bosch, Juergen wrote

Re: [ccp4bb] Resolution, R factors and data quality

2013-08-27 Thread Bosch, Juergen
Hi Jim, all data is good data - the more data you have the better (that's what they say anyhow) Not everybody is adopting to the Karplus Diederich paper as quickly as you do. And not to be confused with the Diederichs and Karplus paper :-) http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3689524/

Re: [ccp4bb] Stuck rfree - possible non merohedral twinning ?

2013-08-25 Thread Bosch, Juergen
Hi Mahesh, if you use Refmac, then you can tell it to refine the twin fraction, no need to tell it the twin law as Refmac will figure it out. If you use phenix, you explicitly tell it the twin law and refine then with it. You can get the possible twin laws by running phenix.xtriage and looking

Re: [ccp4bb] cryoprotection

2013-08-23 Thread Bosch, Juergen
increase your PEG3350 27% and keep the other components at their current concentration. You can also add glycerol or ethylene glycol into the mix. If you have multiple crystals then try various variants. Jürgen On Aug 23, 2013, at 1:52 PM, Uday Kumar wrote: Hello Can anyone suggest a

Re: [ccp4bb] Low 280 absorbance imidazole?

2013-08-22 Thread Bosch, Juergen
Yes, I'm also surprised why people run gradients for the capturing step ? How often is your desired protein spread out throughout the whole gradient ? So what's the point the of the gradient if in the end you merge the whole peak again ? Maybe for a pilot study to identify how much imidazole

Re: [ccp4bb] Low 280 absorbance imidazole?

2013-08-22 Thread Bosch, Juergen
, 2013-08-22 at 10:07 -0400, Bosch, Juergen wrote: Yes, I'm also surprised why people run gradients for the capturing step ? Because we can. Joking aside, I've seen some examples where protein eluted at relatively low imidazole and upon running the gradient there remains some (minimal) overlap

Re: [ccp4bb] Low 280 absorbance imidazole?

2013-08-22 Thread Bosch, Juergen
mashed potatoes On Aug 22, 2013, at 2:17 PM, Ed Pozharski wrote: On Thu, 2013-08-22 at 13:58 -0400, Bosch, Juergen wrote: well if we hit the timer after lysis, say via cell disruptor then I have my eluted protein in less than 1 hour, including 40 minutes batch binding. then proceeds to wait six

Re: [ccp4bb] Low 280 absorbance imidazole?

2013-08-21 Thread Bosch, Juergen
How about low pH elution or EDTA as alternative ? Jürgen .. Jürgen Bosch Johns Hopkins Bloomberg School of Public Health Department of Biochemistry Molecular Biology Johns Hopkins Malaria Research Institute 615 North Wolfe Street, W8708 Baltimore, MD 21205 Phone:

Re: [ccp4bb] Why MAD didn't work but SAD works well

2013-08-20 Thread Bosch, Juergen
Dear George, if #4 is correct, shouldn't he be able to get good SIRAS using the peak dataset as HA and the last collected dataset as native Jürgen .. Jürgen Bosch Johns Hopkins University Bloomberg School of Public Health Department of Biochemistry Molecular Biology Johns

Re: [ccp4bb] non-diffracting xtals

2013-08-19 Thread Bosch, Juergen
Well said Petri, also how much PEG3350 do you have in your conditions ? More than 25% ? I'm going after cryo-conditions at this point, you might want to replace your PEG3350 with smaller PEGs or a mixture of PEG400 and PEG3350. Almost sounds as if no optimization of the original conditions was

Re: [ccp4bb] Data processing - twinned xtals

2013-08-16 Thread Bosch, Juergen
Hi Mahesh, TESTGEN START 1 END 120 ANGLE 0.5 moving the detector further back or using 2theta and collecting more frames. When you started shooting at your image you should have checked for overlaps ahead of time. I assume this to about 1.8 Å - maybe it would have been wiser to collect a 2.5 Å

Re: [ccp4bb] Data processing - twinned xtals

2013-08-16 Thread Bosch, Juergen
for #2) I'd suggest get some of those Mitigen loops that are titled. I assume you have hexagonal plates as crystals and you really want to shoot along the thin area of the crystal down the sixfold. With normal loops it's an art to get that crystal to sit upright in the loop but not impossible

Re: [ccp4bb] Data processing - twinned xtals

2013-08-16 Thread Bosch, Juergen
tilted is what I meant at an angle of e.g. 30 or 60 degrees. Works fine with most SSRL beamlines except of the 12-2 microfocus - but that might have been fixed in the meantime. Jürgen On Aug 16, 2013, at 1:57 PM, Bosch, Juergen wrote: for #2) I'd suggest get some of those Mitigen loops

[ccp4bb] Buccaneer question @Kevin

2013-08-12 Thread Bosch, Juergen
Hi Kevin et al. Buccaneer IS a great program and I like it very much. I have a testcase here for my X-ray class where a trimer is in the asymmetric unit, however all copies are different (not in sequence, but in space - think of Calmodulin for example). There is no option to force Buccaneer

Re: [ccp4bb] Error running Pointless in CCP4-6.3 OSX (10.8)

2013-08-11 Thread Bosch, Juergen
Hi Sheena, simple quickfix to your problem: a) Run CAD and limit the output resolution on that mtz file so you don't run into the memory allocation error of pointless. b) try phenix.xtriage also if it is not a CCP4 program Jürgen .. Jürgen Bosch Johns Hopkins University

Re: [ccp4bb] Substrate/Ligand Induced Oligomerization of enzymes

2013-08-08 Thread Bosch, Juergen
FKBP ? http://www.pnas.org/content/97/13/7096.full.pdf Jürgen On Aug 8, 2013, at 8:03 PM, Shiva Bhowmik wrote: Dear All, I am looking for references and/or example of substrate or ligand induced oligomerization of enzymes related to activation. Any help in this regard would be greatly

Re: [ccp4bb] calculation of shape complementarity of different protein-ligand complexes

2013-08-07 Thread Bosch, Juergen
VROCS, www.eyesopen.comhttp://www.eyesopen.com Jürgen On Aug 7, 2013, at 9:03 AM, Tobias Beck wrote: Dear CCP4bb, I would like to calculate the shape complementarity of several protein-ligand complexes (crystal structures with ligand available). This involves a set of different proteins and

Re: [ccp4bb] Spots not getting indexed properly in protein-DNA complex

2013-08-01 Thread Bosch, Juergen
Hi Appu, nothing really to worry about if you process your data with XDS or d*trek using 3D profile fitting. You still should be able to get something useful out of this data. If you look at 90.png (I'm attaching a zoomed area of your image), you see your crystal is either split or you had two

Re: [ccp4bb] Dose anyone see this ligand before?

2013-07-17 Thread Bosch, Juergen
Try here: http://pubchem.ncbi.nlm.nih.gov/summary/summary.cgi?cid=8031loc=ec_rcs download the sdf and use the sdf to pdb converter Or I think phenix can read in sdf and convert them Jürgen On Jul 17, 2013, at 9:35 AM, Wei Feng wrote: Dear all, Thank you for your advices. I had tried to use MPD

Re: [ccp4bb] Dose anyone see this ligand before?

2013-07-16 Thread Bosch, Juergen
Special position ? sulfate-size ? Jürgen On Jul 16, 2013, at 11:35 AM, Wei Feng wrote: Dear all, I found some redundant density in my structure beween two molecule. (see picture 1 and 2) But I am not sure which ligand will be. Dose everyone see this ligand before? If so, can you tell the PDB

Re: [ccp4bb] Dose anyone see this ligand before?

2013-07-16 Thread Bosch, Juergen
product of reaction. Vaheh From: CCP4 bulletin board [mailto:CCP4BB@JISCMAIL.AC.UK] On Behalf Of Bosch, Juergen Sent: Tuesday, July 16, 2013 11:53 AM To: CCP4BB@JISCMAIL.AC.UKmailto:CCP4BB@JISCMAIL.AC.UK Subject: Re: [ccp4bb] Dose anyone see this ligand

Re: [ccp4bb] Concentrating purified membrane protein

2013-07-14 Thread Bosch, Juergen
can you elute off the column with low pH ? Or EDTA if you don't want imidazole around ? Jürgen On Jul 14, 2013, at 10:29 AM, Raji Edayathumangalam wrote: Thanks everyone for your responses. I definitely plan to save the flowthrough so we'll see what happens. My protein has a His tag and I did

Re: [ccp4bb] XDS not picking spots correctly, please help!

2013-07-04 Thread Bosch, Juergen
Hi Fei, I assume you have edited the XDS.INP file to include the initially found space group and the first refined cell ? If not, then that would be my first choice to do. If that then still fails with the default parameters, then I would start fixing some values instead of refining all of

[ccp4bb] quick off topic question

2013-07-01 Thread Bosch, Juergen
Anybody out there who knows how many people are subscribed to the BB ? Just curious. And no please don't reply all :-) Jürgen .. Jürgen Bosch Johns Hopkins University Bloomberg School of Public Health Department of Biochemistry Molecular Biology Johns Hopkins Malaria

Re: [ccp4bb] Hi

2013-06-10 Thread Bosch, Juergen
Hi Eleanor, C2 - this was XDS lingo or Bravais talk :-) Jürgen ** LATTICE SYMMETRY IMPLICATED BY SPACE GROUP SYMMETRY ** BRAVAIS-POSSIBLE SPACE-GROUPS FOR PROTEIN CRYSTALS TYPE [SPACE GROUP NUMBER,SYMBOL] aP [1,P1] mP [3,P2] [4,P2(1)] mC,mI

Re: [ccp4bb] Hi

2013-06-10 Thread Bosch, Juergen
The other obvious conclusion would be that dataset #3 is a different protein perhaps ? How about pointless for your third dataset ? Jürgen On Jun 10, 2013, at 2:57 PM, Wei Shi wrote: Hi all, I was trying to solve the structure of a protein in several different datasets using xds and phenix. I

Re: [ccp4bb] Helix forming proteins crystallizing as rings

2013-06-05 Thread Bosch, Juergen
Hi Joern, have a look at these two papers (and probably KK is reading this too and can chime in with the right set of coordinates), Korotkov et al. Secretins: dynamic channels for protein transport across membranes. Trends Biochem Sci (2011) vol. 36 (8) pp. 433-43 Reichow et al. Structure of

Re: [ccp4bb] Active site volume calculator

2013-06-05 Thread Bosch, Juergen
Hi Yuri, http://fpocket.sourceforge.net http://sts-fw.bioengr.uic.edu/castp/calculation.php Just to name 2 Jürgen On Jun 5, 2013, at 7:12 PM, Yuri Pompeu wrote: Dear BB, I am sorry for posting off-topic but it is hard not to ask when you know you can get a good answer ;-) I need to

Re: [ccp4bb] visualizing protein-protein interface

2013-06-04 Thread Bosch, Juergen
Hi Tobias, this may or may not work. calculate a map for both proteins, normalize them, put them on an identical cell and subtract one map from another. The program that can do this is called mapman *gosh* a non-CCP4 thing. Good luck, Jürgen On Jun 4, 2013, at 10:18 AM, Tobias Beck wrote:

Re: [ccp4bb] anomalous scattering server down?

2013-06-01 Thread Bosch, Juergen
seems to be down sorry. you should be able to use crossec from ccp4 Jürgen On Jun 1, 2013, at 6:47 PM, Edward A. Berry wrote: Is Ethan Merritt's anomalous scattering page at: http://www.bmsc.washington.edu/scatter/ down or moved, or the firewall I'm behind is blocking it? I want to check

Re: [ccp4bb] how to make the crystal thicker

2013-05-28 Thread Bosch, Juergen
Instead of making the crystal thicker, why don't you make the beamsize smaller ? Aka microfocus If this is your cryo-condition then I would work on it a bit more or change the drop ratio of protein:reservoir so that you can use a higher PEG3350 concentration in your reservoir. I would aim for

Re: [ccp4bb] Organic solvents for ligand solubilisation

2013-05-23 Thread Bosch, Juergen
Hi Klaus, - small molecular weight PEG's e.g. 200 instead of DMSO, has the advantage of also helping to cryo protect - Methanol (only for dispensing the compound into wells) then allow to evaporate and simply add your cryo-protected crystals, the hope is that sufficient of your ligand goes

Re: [ccp4bb] cryocrystals

2013-05-22 Thread Bosch, Juergen
I dare say indefinitely. Your crystal life expectancy drops rapidly though with somebody not remembering to fill up the dewar. Transferring crystals to pucks etc. is another problem. I had some crystals which I could not remember what they were (label broke off the cane) and we simply reshoot

Re: [ccp4bb] how to solve crystal structure using published EM structure

2013-05-18 Thread Bosch, Juergen
You can use it as an MR model in Molrep for example. Or are you asking where to get the actual EM data from ? That would be here: http://www.ebi.ac.uk/pdbe/emdb/ Jürgen On May 18, 2013, at 8:32 PM, LISA wrote: Hi All, The EM structure of a complex was published at 8A. If we can collect

Re: [ccp4bb] disordered helix

2013-05-14 Thread Bosch, Juergen
This looks to me like a good attempt for GraphEnt. Give it a shot and you might be positively surprised. Jürgen Sent from my iPad On May 13, 2013, at 21:26, Dale Tronrud det...@uoxray.uoregon.edu wrote: Sometimes a floppy bit of a protein is even more floppy in a particular crystal form.

Re: [ccp4bb] off topic question regarding SPR

2013-05-05 Thread Bosch, Juergen
Hi Faisal, whatever makes your protein happy is good. What type of experiment were you thinking of ? Protein-protein interaction or protein-small molecule interaction ? In the latter case add some Tween20 or Brij-35 into the mix. A decent overview can be found under this webpage for what to

Re: [ccp4bb] need suggestions

2013-04-16 Thread Bosch, Juergen
Hi Afshan, You mention: The protein prep is same which was using to get the Hits. So how old is your prep ? Was it stored at 4C or in LN2? Do you know if your protein is stable under the conditions you stored it ? How long after purification did it take you to get those crystals ? How old is

Re: [ccp4bb] 2Theta data set solving problem

2013-04-09 Thread Bosch, Juergen
I thought imosflm recognizes automatically the 2theta offset ? Jürgen .. Jürgen Bosch Johns Hopkins Bloomberg School of Public Health Department of Biochemistry Molecular Biology Johns Hopkins Malaria Research Institute 615 North Wolfe Street, W8708 Baltimore, MD 21205

Re: [ccp4bb] delete subject

2013-03-29 Thread Bosch, Juergen
There's a second side to that. Reviewers who can't get enough data and request even more when you submit a decent paper with 18 pages of supplement for example. Jürgen On Mar 29, 2013, at 7:32 AM, Toufic El Arnaout wrote: http://www.plosone.org/article/info:doi%2F10.1371%2Fjournal.pone.0053374

Re: [ccp4bb] refinement protein structure

2013-03-27 Thread Bosch, Juergen
Hi Tom, some suggestions for you: You should calculate a selfrotation function and see if you can learn something from it. You should definitely run Matthews Coefficient and see if you get a brilliant idea there And congratulations to your first crystal structure it looks really great, for a

Re: [ccp4bb] Twinning problem

2013-03-26 Thread Bosch, Juergen
get the twin law and either refine with phenix.refine twin_law=-h,-k,l or whatever it suggests, or just add into your Refmac script the line TWIN and it will figure out the twin law for you. You can also detwin data but then you might be throwing away a lot of data. We've now had two cases

Re: [ccp4bb] Isothermal titration calorimetry

2013-03-24 Thread Bosch, Juergen
Hi George, this is probably a very stupid suggestion and you likely have tried it, but I'll suggest the obvious nevertheless. What happens to your .nitc file when you rename it to .itc can you read it in Origin then ? Jürgen On Mar 24, 2013, at 6:39 AM, George Kontopidis wrote: Chris,

Re: [ccp4bb] ionic interaction inside a protein

2013-03-23 Thread Bosch, Juergen
Try PDBSum Jürgen On Mar 23, 2013, at 9:34 AM, Faisal Tarique wrote: Dear all I am working on a thermostable protein and i have read that the stability to high temperature is due to various ionic interactions among the amino acid residues of the protein itself..I request you all to tell me

Re: [ccp4bb] Map Alignment

2013-03-21 Thread Bosch, Juergen
If I understand your question correctly, you have a couple of atoms which you could align to get the rotation translation then you can use these values with maprot (CCP4) or mama (USF) to actually superimpose maps. Jürgen On Mar 21, 2013, at 11:29 PM, Chen Zhao wrote: Dear all, Does anybody

Re: [ccp4bb] suitable buffer for CD studies

2013-03-20 Thread Bosch, Juergen
Why not thermal denaturation in the presence of Sypro Orange and a realtime PCR machine ? Crowther et al. Use of thermal melt curves to assess the quality of enzyme preparations. Anal Biochem (2010) vol. 399 (2) pp. 268-275 Or (shameless advertisement): Hain et al. Structural characterization

Re: [ccp4bb] suitable buffer for CD studies

2013-03-20 Thread Bosch, Juergen
Yep, mostly you should stay away from Tris as this is the worst buffer system when playing with temperature changes. Tris for example has a ∆pKa/10˚C -0.31 Good, N.E. (1986) Biochemistry 5, 467 Jürgen P.S. @Matthew, was this what you meant by the Good buffers often not ? or just a

Re: [ccp4bb] first use of synchrotron radiation in PX

2013-03-16 Thread Bosch, Juergen
Thank you James, you should write a History book about the modern x-ray times. Or better make one of those movies you are famous for. Jürgen On Mar 16, 2013, at 10:46 AM, James Holton wrote: The first report of shooting a protein crystal at a synchrotron (I think) was in 1976:

Re: [ccp4bb] [ccp4bb] validating ligand density

2013-03-12 Thread Bosch, Juergen
Going back to the initial question. I would recommend looking at AFITT http://www.eyesopen.com/afitt Works like a dream (in certain cases). Jürgen P.S. I wish I had some stocks from them but I don't .. Jürgen Bosch Johns Hopkins University Bloomberg School of Public Health

Re: [ccp4bb] How to compare B-factors between structures?

2013-03-04 Thread Bosch, Juergen
Yep, I agree calculate the average B per structure and divide each B by this value, then multiply it by any value that is reasonable so you can visualize color differences :-) Jürgen On Mar 4, 2013, at 2:16 PM, Jacob Keller wrote: You only entertain addition+subtraction--why not use

Re: [ccp4bb] B factor of the loop

2013-03-03 Thread Bosch, Juergen
yes - but keep in mind your protein is in the context of the crystal lattice, so flexible regions in solution are likely to be stabilized in the crystal lattice. So if you color by B also look at the symmetry mates. And you should also submit both structures to the TLSMD server and look at

Re: [ccp4bb] off topic question BIAcore problem

2013-02-20 Thread Bosch, Juergen
I assume you use CM5 chips ? I further assume you run at pH 7.5 perhaps ? What's the pI of your analyte ? 7.5 ? Do you get significant binding to your reference cell under the conditions you are running ? You might get rebinding to your negatively charged surface and the dissociation you are

Re: [ccp4bb] off topic question BIAcore problem

2013-02-20 Thread Bosch, Juergen
Well that looks pretty real then. You might have wrong concentrations in one or the other experiment perhaps hence the difference. Jürgen On Feb 20, 2013, at 5:45 PM, xianchi dong wrote: Thanks a lot for the kind reply. I used CM5 chip at pH 7.4. I am not quite sure about the PI of my protein

Re: [ccp4bb] Mac mini advice

2013-01-23 Thread Bosch, Juergen
I assume nobody of you is running an actual Osx server ? I mean the upgrade to a full server version of the commonly distributed normal Osx releases ? I have not done it yet but I do think many of the issues mentioned regarding NFS/NIS could be addressed there. Regarding the missing macpro

Re: [ccp4bb] Mac mini advice

2013-01-22 Thread Bosch, Juergen
Current Mac minis outperform my 2009 models with which I am still happy. So dual core I guess would be sufficient no need for upgrade on graphics card. Jürgen .. Jürgen Bosch Johns Hopkins Bloomberg School of Public Health Department of Biochemistry Molecular Biology Johns

Re: [ccp4bb] Mac mini advice

2013-01-22 Thread Bosch, Juergen
Any reason for the Mac Mini over the iMac A zalman monitor ir can you hook up a second monitor in stereo mode to your iMac ? Jürgen .. Jürgen Bosch Johns Hopkins Bloomberg School of Public Health Department of Biochemistry Molecular Biology Johns Hopkins Malaria Research

Re: [ccp4bb] a challenge

2013-01-14 Thread Bosch, Juergen
What is the best procedure to use for weak anomalous signal That opens up the can of worms which I'm happy to jump into. We've had very good success in the years 2003-2009 with shelx for finding sites (sometimes more than 1 trials) then force feeding them to sharp for phase improvement. We

Re: [ccp4bb] Coot's hidden talents!

2013-01-11 Thread Bosch, Juergen
Surprised Mercedes didn't sued him for that :-) @Mr. Emsley I assume you soon will be Sir Emsley after that marvelous painting.mentioned in Harry's email. Jürgen .. Jürgen Bosch Johns Hopkins University Bloomberg School of Public Health Department of Biochemistry Molecular

Re: [ccp4bb] Symmetry operator

2013-01-10 Thread Bosch, Juergen
coot: show symmetry molecules, then save symmetry molecule and you have your tetramer most likely Jürgen On Jan 10, 2013, at 7:48 PM, james09 pruza wrote: Hi, Which program outputs the symmetry operator (rotation and translation)? I have a dimer in the asymmetric unit and need to know the

Re: [ccp4bb] Who invented PDB format?

2013-01-06 Thread Bosch, Juergen
bash or any other shell script using http://www.ccp4.ac.uk/html/coordconv.html Probably moleman2 http://xray.bmc.uu.se/usf/moleman_man.html can handle your request too. Jürgen On Jan 7, 2013, at 12:48 AM, Teri Arman wrote: Fractional Coordiantes to Orthogonal Coordinates and Vice Versa Hi, I

Re: [ccp4bb] 3D alignment of points (atoms)

2012-12-28 Thread Bosch, Juergen
can't you feed solve with those positions or Hyss or the Sharp interface ? It's been too long ago when I faced this issue but I remember force-feeding one of the existing programs at the time with blank coordinates to figure it out - or was it mlphare ? Too long ago probably 10 years by now.

Re: [ccp4bb] Today ...

2012-12-21 Thread Bosch, Juergen
the months renamed alphabetically. On 12/21/12 03:23, Tom Murray-Rust wrote: Hi Juergen, Your scheme as printed has two J's - so January and July are indistinguishable! I would suggest the letters should instead be JFMAYULGSOND. On 21 Dec 2012, at 01:52, Bosch, Juergen jubo

Re: [ccp4bb] archival memory?

2012-12-12 Thread Bosch, Juergen
Hey Dale, you really should get your personal RAID with hot swappable discs, since you don't like Firewire, how about Thunderbolt and a Pegasus RAID with 6 bays ? If a drive fails you replace it with a new one. By the way if anybody has a functional DAT4 tape drive, could I send you one to

Re: [ccp4bb] Collecting native data at an absorption edge

2012-11-20 Thread Bosch, Juergen
You'll see the zinc also at the Semet peak edge already check out Ethan's edge plot web server. If you used his-tag purification methods it would be wiser to collect passed the edge of either Ni or Co so you can distinguish them from Zn sites. Jürgen .. Jürgen Bosch Johns

Re: [ccp4bb] OT: Has anyone experienced problems with Apple laptop battery expansion?

2012-11-20 Thread Bosch, Juergen
It probably had a functional Firewall under Linux that's why it survived :-) Jürgen P.S. note to myself, don't ever give George my laptop, as it might be mistreated - just for test purposes :-) On Nov 20, 2012, at 12:57 PM, George M. Sheldrick wrote: It is dual bootable Linux/Windows George

Re: [ccp4bb] Off topic: Selecting atoms within a given distance from a target atom

2012-11-18 Thread Bosch, Juergen
Hi Pavel, does this also work for symmetry related atoms ? Jürgen .. Jürgen Bosch Johns Hopkins Bloomberg School of Public Health Department of Biochemistry Molecular Biology Johns Hopkins Malaria Research Institute 615 North Wolfe Street, W8708 Baltimore, MD 21205 Phone:

Re: [ccp4bb] OT: Has anyone experienced problems with Apple laptop battery expansion?

2012-11-18 Thread Bosch, Juergen
Bill I think that's crap. I had issues on a 2005 MacBook Pro with inflating battery and it was replaced (after about 6 months). There were troubles with those batteries and impurities but mine still had apple care at that time and the batteries were exchangeable. I have not heard of the build

Re: [ccp4bb] vitrification vs freezing

2012-11-16 Thread Bosch, Juergen
HI Tim, you should know better. German is the most precise language, hence all those old German *gosh* books (for the younger readers of this board, there was a time before pdf and Nook readers) for organic chemistry etc. from the 19th century and older (Beilstein, Angewandte ...). And why was

Re: [ccp4bb] off topic: oligomer detection

2012-11-14 Thread Bosch, Juergen
What if your mild urea condition unfolds your protein from a globular to a stretched elongated form ? You probably have done CD already to verify that this is not the case. Jürgen .. Jürgen Bosch Johns Hopkins Bloomberg School of Public Health Department of Biochemistry

Re: [ccp4bb] Apologies

2012-11-12 Thread Bosch, Juergen
Hi Matthias, I have not read your attached MS however Figure 2 is very good. Congrats to the first author for a very captive and engaging view of your protein. Jürgen On Nov 12, 2012, at 4:24 AM, Matthias Wilmanns wrote: I am apologizing for sending an unintended email to CCP4 via CC.

Re: [ccp4bb] usefulness of cacodylate?

2012-11-09 Thread Bosch, Juergen
Well originally we got these in cacodylate and not much else would diffract. http://www.pdb.org/pdb/explore/materialsAndMethods.do?structureId=2EPH But nowadays I have another conditions, which does not require cacodylate and works well with Hepes. Jürgen On Nov 9, 2012, at 7:26 AM, Frank von

Re: [ccp4bb] What to put on Custom Declaration for shipped samples?

2012-11-07 Thread Bosch, Juergen
Hi Frank, I like your letter with the random initiative :-) And then the thread by citing a regulation, which they probably never have heard of. I would additionally add the regulation numbers for the dewar. There are two or three IATA numbers that should be mentioned to indicate its in line

Re: [ccp4bb] protein cleavage

2012-11-04 Thread Bosch, Juergen
@Cynthia, On Nov 4, 2012, at 10:58 AM, Cynthia Kinsland wrote: Since you're seeing the MBP band, it sounds as if you're getting some cleavage. If there were no cleave you would only see the fusion and TEV bands on the gel. I think that is a wrong assumption. He did not specify if he sees the

Re: [ccp4bb] Ca or Zn

2012-10-30 Thread Bosch, Juergen
calculate an anomalous map, you should see the Zn signal even if you collected at the SeMet peak. Jürgen .. Jürgen Bosch Johns Hopkins University Bloomberg School of Public Health Department of Biochemistry Molecular Biology Johns Hopkins Malaria Research Institute 615 North

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