[ccp4bb] Off topic, but am desparate...Ah, a solution!

[Jacob Keller]

Hi All,

Thanks for all of the great suggestions--it was actually Ethan Merritt's 
suggestions and pointing to the right Gnuplot documentation that really 
worked for me. Gnuplot is actually pretty powerful, and can make quite nice 
images. Again, thanks a million for the help.


Jacob Keller

- Original Message - 
From: "Ethan Merritt" <[EMAIL PROTECTED]>

To: 
Sent: Wednesday, January 16, 2008 5:19 PM
Subject: Re: [ccp4bb] Off topic, but am desparate...



On Wednesday 16 January 2008 13:27, Jacob Keller wrote:

Hello all,

It might just take you a few minutes to tell me how to do this:

I have a set of data in three columns xyz (resid1, resid2, value), which 
I
would like to plot as a "heat" or "color map" on the 2d rectangle formed 
by
the values xy, and colored by z. Essentially what I want is a ~1500 x 
1500
grid with boxes color-coded by their z values. I cannot figure out how to 
do

this in gnuplot or anywhere, despite efforts to learn gnuplot and other
programs. Any suggestions?


Use gnuplot.

If you have complete data, i.e. every pair [x,y] is represented,
then the gnuplot command script is very simple so long as the
data file format is correct.  Two formats are possible.
Here is the one closest to what you describe.

file:
0 0 f(0,0)
0 1 f(0,1)
...
0 n f(0,n)
   blank line.  Very important!
1 0 f(1,0)
1 1 f(1,1)
...
1 n f(1,n)
   blank line.  Very important!
2 0 f(2,0)
... and so on


gnuplot commands:
  # You must supply actual values for min/max
  # because image mode does not autoscale
  set xrange [zmin:xmax]
  set yrange [ymin:ymax]
  set cbrange [zmin:zmax]
  plot 'data' using 1:2:3 with image


The other option is to reformat your data into a file containing
only the z values.  [x,y] is implicit from the location in the file:

file:

f(0,0) f(0,1) ... f(0,n)
f(1,0) f(1,1) ... f(1,n)
...
f(n,0) f(n,1) ... f(n,n)

gnuplot commands:
  # this time we use a 3D command 'splot'
  set view map
  splot 'data' matrix using 1:2:3 with image

If the data is sparse, then it's still possible but the command
sequence is more complicated and it uses "with pm3d" to interpolate
a surface rather than "with image" with treats it as a complete set
of pixels.

More gnuplot sample scripts online at
   http://gnuplot.sourceforge.net/demo/

Ethan

--
Ethan A MerrittCourier Deliveries: 1959 NE Pacific
Dept of Biochemistry
Health Sciences Building
University of Washington - Seattle WA 98195-7742

Re: [ccp4bb] crystallisation robot

[mjvdwoerd]

 Once upon a time I worked in a group that was interested in developing 
crystallization in microfluidics. This was before the time that Fluidigm 
existed and we had not heard of crystallization with the aid of microfluidics 
at the time. We had good reason to try to make a system that was as small and 
light as possible - it had something to do with the cost of shipping proteins 
and precipitants - less was better. And we also wanted all protein drops to be 
fully enclosed, out of safety considerations.

Like Tassos, we were very worried what would happen if you scaled back drops 
along the lines of this discussion - several uL downto tens of nanoliters. If 
the stochastic process had a major influence over this process, we thought that 
we would never get any crystals. So we set up side-by-side experiments at 
larger volumes and smaller volumes - basically scanning several orders of 
magnitude - expecting a decrease of the number of crystals when volumes 
decrease. To our great surprise the outcome was that smaller volumes almost 
always gave MORE (I almost want to say 'dramatically more') crystals, more 
nucleation, and indeed in various cases the crystals grew much faster also. 
Indeed, it was trivial to observe that the surface-to-volume ratio was the 
primary driver for the nucleation process. We had control over geometry to some 
extent and were able to observe surfaces while crystals grow. The crystals 
would most commonly nucleate on a surface. 

So although there probably is something to stochastic aspects, it is clear that 
other aspects can be more important and "overrule" the stochastic 
considerations.
The somewhat unpleasant consquence is of course that results acquired in very 
small volumes (with larger surface-to-volume ratio) cannot necessarily be 
repeated in larger volumes (smaller surface-to-volume ratio).

This is not a flame, even if heat might be a good thing on a night with 
temperatures predicted far below 0F.

?:-)

Mark


 


 

-Original Message-
From: Anastassis Perrakis <[EMAIL PROTECTED]>
To: CCP4BB@JISCMAIL.AC.UK
Sent: Wed, 16 Jan 2008 6:17 am
Subject: Re: [ccp4bb] crystallisation robot









> Oryxnano 50+50 nL?

>?

> Demetres?

>?
?

Which, indirectly, brings up an interesting (but not relevant to the Oryx) 
question.?
?

Nucleation is a process that does have a stochastic aspect.?
?

Thus, one could argue that compromising to 200-300 nl might be better than 
either extremes of 50nl (too small volume and less chance for nucleation) or 
1000 nl (too much sample).?
?

any comments ? (let the flames begin).?
?

A.?
?

PS1?

another interesting issue that has has been hardly touched in these emails is 
the real sample loss: left in wells and not easy to recover, lost because of 
contamination with system liquid, etc ...?
?

PS2?

I see lots of people with new robots. please do have a look at the 
www.BIOXHIT.org page and if you have a few minutes to assemble a table we will 
be happy to add your specs to our pages. it can be a nice resource and it has 
already enough things and already one response to my last email ;-) To make 
life easier to potential contributors we can provide an Excel sheet to fill up 
with your specs - just ask.?
?

On Jan 16, 2008, at 12:46, Demetres D. Leonidas wrote:?
?

>?

> David Briggs wrote:?

>> I'll defend the honour of the phoenix... (again)?

>>?

>> Bernhard Rupp 100+100 nl?

>> Dave Briggs (and all users at Univ of Manchester, UK) 100+100nl?

>> Others..?

>>?

>> Only time we have ANY problems is when the nano dispensing tip >> gets 
>> clogged. Often a good wash whilst still on the machine will >> clear the 
>> blockage.?

>>?

>> Dave?

>>?

>>?

>>?

>>?

>> -->> ?

>> David C. Briggs PhD?

>> Father & Crystallographer?

>> http://www.dbriggs.talktalk.net ?

>> AIM ID: dbassophile?

>> ?

>?

> --> Demetres D. Leonidas, Ph.D.?

> Structural Biology & Chemistry Group?

> Institute of Organic and Pharmaceutical Chemistry?

> The National Hellenic Research Foundation?

> 48, Vassileos Constantinou Avenue?

> Athens 116 35, Greece?

> ==?

> Tel. +30 210 7273841 (office)?

> +30 210 7273895 (lab) Fax. +30 210 7273831?

> E-mail: [EMAIL PROTECTED]

> URL: http://athena.eie.gr?

> ==?



 



More new features than ever.  Check out the new AIM(R) Mail ! - 
http://webmail.aim.com

Re: [ccp4bb] crystallisation robot

[mjvdwoerd]

 Once upon a time I worked in a group that was interested in developing 
crystallization in microfluidics. This was before the time that Fluidigm 
existed and we had not heard of crystallization with the aid of microfluidics 
at the time. We had good reason to try to make a system that was as small and 
light as possible - it had something to do with the cost of shipping proteins 
and precipitants - less was better. And we also wanted all protein drops to be 
fully enclosed, out of safety considerations.

Like Tassos, we were very worried what would happen if you scaled back drops 
along the lines of this discussion - several uL downto tens of nanoliters. If 
the stochastic process had a major influence over this process, we thought that 
we would never get any crystals. So we set up side-by-side experiments at 
larger volumes and smaller volumes - basically scanning several orders of 
magnitude - expecting a decrease of the number of crystals when volumes 
decrease. To our great surprise the outcome was that smaller volumes almost 
always gave MORE (I almost want to say 'dramatically more') crystals, more 
nucleation, and indeed in various cases the crystals grew much faster also. 
Indeed, it was trivial to observe that the surface-to-volume ratio was the 
primary driver for the nucleation process. We had control over geometry to some 
extent and were able to observe surfaces while crystals grow. The crystals 
would most commonly nucleate on a surface. 

So although there probably is something to stochastic aspects, it is clear that 
other aspects can be more important and "overrule" the stochastic 
considerations.
The somewhat unpleasant consquence is of course that results acquired in very 
small volumes (with larger surface-to-volume ratio) cannot necessarily be 
repeated in larger volumes (smaller surface-to-volume ratio).

This is not a flame, even if heat might be a good thing on a night with 
temperatures predicted far below 0F.

?:-)

Mark


 


 

-Original Message-
From: Anastassis Perrakis <[EMAIL PROTECTED]>
To: CCP4BB@JISCMAIL.AC.UK
Sent: Wed, 16 Jan 2008 6:17 am
Subject: Re: [ccp4bb] crystallisation robot









> Oryxnano 50+50 nL?

>?

> Demetres?

>?
?

Which, indirectly, brings up an interesting (but not relevant to the Oryx) 
question.?
?

Nucleation is a process that does have a stochastic aspect.?
?

Thus, one could argue that compromising to 200-300 nl might be better than 
either extremes of 50nl (too small volume and less chance for nucleation) or 
1000 nl (too much sample).?
?

any comments ? (let the flames begin).?
?

A.?
?

PS1?

another interesting issue that has has been hardly touched in these emails is 
the real sample loss: left in wells and not easy to recover, lost because of 
contamination with system liquid, etc ...?
?

PS2?

I see lots of people with new robots. please do have a look at the 
www.BIOXHIT.org page and if you have a few minutes to assemble a table we will 
be happy to add your specs to our pages. it can be a nice resource and it has 
already enough things and already one response to my last email ;-) To make 
life easier to potential contributors we can provide an Excel sheet to fill up 
with your specs - just ask.?
?

On Jan 16, 2008, at 12:46, Demetres D. Leonidas wrote:?
?

>?

> David Briggs wrote:?

>> I'll defend the honour of the phoenix... (again)?

>>?

>> Bernhard Rupp 100+100 nl?

>> Dave Briggs (and all users at Univ of Manchester, UK) 100+100nl?

>> Others..?

>>?

>> Only time we have ANY problems is when the nano dispensing tip >> gets 
>> clogged. Often a good wash whilst still on the machine will >> clear the 
>> blockage.?

>>?

>> Dave?

>>?

>>?

>>?

>>?

>> -->> ?

>> David C. Briggs PhD?

>> Father & Crystallographer?

>> http://www.dbriggs.talktalk.net ?

>> AIM ID: dbassophile?

>> ?

>?

> --> Demetres D. Leonidas, Ph.D.?

> Structural Biology & Chemistry Group?

> Institute of Organic and Pharmaceutical Chemistry?

> The National Hellenic Research Foundation?

> 48, Vassileos Constantinou Avenue?

> Athens 116 35, Greece?

> ==?

> Tel. +30 210 7273841 (office)?

> +30 210 7273895 (lab) Fax. +30 210 7273831?

> E-mail: [EMAIL PROTECTED]

> URL: http://athena.eie.gr?

> ==?



 



More new features than ever.  Check out the new AIM(R) Mail ! - 
http://webmail.aim.com

[ccp4bb] fffear/pirate/buccaneer and SIGFP

[Bryan W. Lepore]

do any of fffear/pirate/buccaneer use SIGFP in a significant/meaningful 
way?


-bryan

Re: [ccp4bb] Off topic, but am desparate...

[Ethan Merritt]

On Wednesday 16 January 2008 13:27, Jacob Keller wrote:
> Hello all,
> 
> It might just take you a few minutes to tell me how to do this:
> 
> I have a set of data in three columns xyz (resid1, resid2, value), which I 
> would like to plot as a "heat" or "color map" on the 2d rectangle formed by 
> the values xy, and colored by z. Essentially what I want is a ~1500 x 1500 
> grid with boxes color-coded by their z values. I cannot figure out how to do 
> this in gnuplot or anywhere, despite efforts to learn gnuplot and other 
> programs. Any suggestions?

Use gnuplot.

If you have complete data, i.e. every pair [x,y] is represented,
then the gnuplot command script is very simple so long as the
data file format is correct.  Two formats are possible.
Here is the one closest to what you describe.

file:
0 0 f(0,0)
0 1 f(0,1)
...
0 n f(0,n)
    blank line.  Very important!
1 0 f(1,0)
1 1 f(1,1)
...
1 n f(1,n)
    blank line.  Very important!
2 0 f(2,0)
... and so on


gnuplot commands:
   # You must supply actual values for min/max
   # because image mode does not autoscale
   set xrange [zmin:xmax]
   set yrange [ymin:ymax]
   set cbrange [zmin:zmax]
   plot 'data' using 1:2:3 with image


The other option is to reformat your data into a file containing
only the z values.  [x,y] is implicit from the location in the file:

file:

f(0,0) f(0,1) ... f(0,n)
f(1,0) f(1,1) ... f(1,n)
...
f(n,0) f(n,1) ... f(n,n)

gnuplot commands:
   # this time we use a 3D command 'splot'
   set view map
   splot 'data' matrix using 1:2:3 with image

If the data is sparse, then it's still possible but the command
sequence is more complicated and it uses "with pm3d" to interpolate
a surface rather than "with image" with treats it as a complete set
of pixels.

More gnuplot sample scripts online at
http://gnuplot.sourceforge.net/demo/

Ethan

-- 
Ethan A MerrittCourier Deliveries: 1959 NE Pacific
Dept of Biochemistry
Health Sciences Building
University of Washington - Seattle WA 98195-7742

Re: [ccp4bb] Off topic, but am desparate...

[Bryan W. Lepore]

On Wed, 16 Jan 2008, Jacob Keller wrote:
I would like to plot as a "heat" or "color map" on the 2d rectangle 
[...] cannot figure out how to do this in gnuplot or anywhere,


consider gnuplot pm3d (not readily obvious for googling, is it)

http://t16web.lanl.gov/Kawano/gnuplot/plotpm3d-e.html

-bryan

Re: [ccp4bb] Off topic, but am desparate...

[Tim Gruene]

Are you familiar with pm3d? I couldn't give the solution from the top of 
my head, but

http://t16web.lanl.gov/Kawano/gnuplot/plotpm3d-e.html
seems a good source of information.


--
Tim Gruene
Institut fuer anorganische Chemie
Tammannstr. 4
D-37077 Goettingen

GPG Key ID = A46BEE1A


On Wed, 16 Jan 2008, Jacob Keller wrote:


Hello all,

It might just take you a few minutes to tell me how to do this:

I have a set of data in three columns xyz (resid1, resid2, value), which I 
would like to plot as a "heat" or "color map" on the 2d rectangle formed by 
the values xy, and colored by z. Essentially what I want is a ~1500 x 1500 
grid with boxes color-coded by their z values. I cannot figure out how to do 
this in gnuplot or anywhere, despite efforts to learn gnuplot and other 
programs. Any suggestions?


Best regards,

Jacob Keller

***
Jacob Pearson Keller
Northwestern University
Medical Scientist Training Program
Dallos Laboratory
F. Searle 1-240
2240 Campus Drive
Evanston IL 60208
lab: 847.491.2438
cel: 773.608.9185
email: [EMAIL PROTECTED]
***

[ccp4bb] Off topic, but am desparate...

[Jacob Keller]

Hello all,

It might just take you a few minutes to tell me how to do this:

I have a set of data in three columns xyz (resid1, resid2, value), which I 
would like to plot as a "heat" or "color map" on the 2d rectangle formed by 
the values xy, and colored by z. Essentially what I want is a ~1500 x 1500 
grid with boxes color-coded by their z values. I cannot figure out how to do 
this in gnuplot or anywhere, despite efforts to learn gnuplot and other 
programs. Any suggestions?


Best regards,

Jacob Keller

***
Jacob Pearson Keller
Northwestern University
Medical Scientist Training Program
Dallos Laboratory
F. Searle 1-240
2240 Campus Drive
Evanston IL 60208
lab: 847.491.2438
cel: 773.608.9185
email: [EMAIL PROTECTED]
***

Re: [ccp4bb] Potassium Nitrate

[Roger Rowlett]

sajid akthar wrote:


Hi All

 

Sorry for non CCP4 question. I want to make Pottasium Nitrate solution 
of pH 6.9 (ofcourse for crystallisation trails only). I'm adjusting pH 
by using NaOH. But it seems the pH is not stable after sometime. Is 
there any good way to do it.


 


Thank you

 


Sajid



Now you can chat without downloading messenger. Click here 
 
to know how.
I would think a solution potassium nitrate in deionized water would have 
a negligible effect on the pH of a buffered crystallization solution. I 
wouldn't bother trying to adjust the pH, because the solution is not 
sufficiently buffered, and depending on the ionic strength of the 
solution, may not have sufficient conductivity to allow a pH electrode 
to work properly.


Cheers,


--

Roger S. Rowlett
Professor
Colgate University Presidential Scholar
Department of Chemistry
Colgate University
13 Oak Drive
Hamilton, NY 13346

tel: (315)-228-7245
ofc: (315)-228-7395
fax: (315)-228-7935
email: [EMAIL PROTECTED]

[ccp4bb] Potassium Nitrate

[sajid akthar]

Hi All

Sorry for non CCP4 question. I want to make Pottasium Nitrate solution of pH 
6.9 (ofcourse for crystallisation trails only). I'm adjusting pH by using NaOH. 
But it seems the pH is not stable after sometime. Is there any good way to do 
it.

Thank you

Sajid


  Now you can chat without downloading messenger. Go to 
http://in.messenger.yahoo.com/webmessengerpromo.php

Re: [ccp4bb] How to improve the density of another molecule in Asymmetric unit

[Lijun Liu]

First you cannot normally expect two in the same AU have the same  
structure, so an over-weighted averaging may cause bias.


It is likely that a rigid-body refinement down to two molecules or  
even domains may help improve in your case.


On Jan 16, 2008, at 8:47 AM, Sun Tang wrote:


Hello Everyone,

I have a question about how to improve the electron density. I have  
two indepedent molecules in the asymmetric unit (1.9 A resolution).  
I solved the structure with PHASER and refined with CCP4i (Rfree =  
0.29).


For the first molecule, the density is very good, but for the  
second one, the density is much worse than the first one. Are there  
any ways to improve the density of the second molecule, such as  
some kinds of averaging? Why that happens?


Thank you very much for your help!

Sincerely,

Sun Tang
[EMAIL PROTECTED]

Never miss a thing. Make Yahoo your homepage.


Lijun Liu, PhD
Institute of Molecular Biology
HHMI & Department of Physics
University of Oregon
Eugene, OR 97403
541-346-4080

Re: [ccp4bb] How to improve the density of another molecule in Asymmetric unit

[Gerard DVD Kleywegt]

ni hao,

 I have a question about how to improve the electron density. I have two 
indepedent molecules in the asymmetric unit (1.9 A resolution). I solved the 
structure with PHASER and refined with CCP4i (Rfree = 0.29).


 For the first molecule, the density is very good, but for the second one, 
the density is much worse than the first one. Are there any ways to improve 
the density of the second molecule, such as some kinds of averaging? Why 
that happens?


there can be various causes for this such as undetected twinning or a 
spacegroup error. if you can exclude those possibilities, it could also be 
that the weak-density molecule has fewer packing interactions and thus is less 
well ordered, and therefore has a higher overall temperature factor. you can 
investigate this with SPANCSI (http://xray.bmc.uu.se/usf/spancsi_man.html#H5).


--dvd

**
Gerard J.  Kleywegt
[Research Fellow of the Royal  Swedish Academy of Sciences]
Dept. of Cell & Molecular Biology  University of Uppsala
Biomedical Centre  Box 596
SE-751 24 Uppsala  SWEDEN

http://xray.bmc.uu.se/gerard/  mailto:[EMAIL PROTECTED]
**
   The opinions in this message are fictional.  Any similarity
   to actual opinions, living or dead, is purely coincidental.
**

[ccp4bb] postdoc position in structural biology of wound healing

[David Hulmes]

STRUCTURAL BIOLOGY OF WOUND HEALING

A 2 year postdoctoral position, funded by the French National Research
Agency, is available at the Institut de Biologie et Chimie des Protéines in
Lyon (www.ibcp.fr) to study the molecular mechanisms of wound healing and to
develop novel anti-scarring strategies. The work will focus on the functions
of the increasingly important family of tolloid metalloproteinases, and
associated proteins, involved in several aspects of tissue repair (see below
for further information).

Candidates should have a strong background in structural biology.

Net salary (after insurance deductions) : 2039 euros/month.

Start date flexible.

To apply, send full CV, list of publications and names and contact details
of at least two scientific referees to:

David Hulmes
Institut de Biologie et Chimie des Protéines
7 passage du Vercors
69367 Lyon cedex 7

Tel : +33 472722667
Fax : +33 472722604
e-mail : [EMAIL PROTECTED]

…

Further information

Research project - The project is funded by the French National Research
Agency. It involves 3 research groups, two based in Lyon and the third in
Toulouse. The idea is to study the role of tolloid proteinases and
associated proteins in the process of wound healing, and to develop novel
anti-scarring strategies. Scarring frequently accompanies wound repair
following events such as surgery, burns and infections. Several structural
proteins, enzymes and growth factors are involved in wound repair. It has
recently become apparent that members of the tolloid family of
metallo-proteinases play several important roles in wound healing and
development (Ge and Greenspan, 2006), either by processing structural
proteins or by activating enzymes and growth factors in the extracellular
matrix (ECM). The activities of tolloid proteinases can be stimulated
several fold by other ECM proteins, called PCPEs. We have recently shown
that PCPE-1 can stimulate the activity of one of the tolloid proteinase
BMP-1, in a substrate specific manner, to selectively accelerate procollagen
processing (Moali et al., 2005) and we have identified structural features
in PCPE-1 required for stimulating activity (Blanc et al., 2007). The aims
of the project are to determine the 3D structures of PCPEs and their
interacting partners in order to develop novel inhibitors to be tested in
both an animal model and using reconstructed corneas.

Blanc G, Font B, Eichenberger D, Moreau C, Ricard-Blum S, Hulmes DJ, Moali C
(2007) J Biol Chem 282, 16924-16933.
Ge G, Greenspan DS (2006) Birth Defects Res C Embryo Today 78: 47-68.
Moali C, Font B, Ruggiero F, Eichenberger D, Rousselle P, Francois V,
Oldberg A, Bruckner-Tuderman L, Hulmes DJS (2005) J Biol Chem 280: 24188-24194.

Working environment - The research will be carried out in the group of David
Hulmes, in collaboration with the group of Richard Haser, at the Institut de
Biologie et Chimie des Protéines (IBCP), which is a joint CNRS/ University
of Lyon institute located in the Gerland region of Lyon. The institute
(www.ibcp.fr) consists of 160 staff (including postdocs and students),
organised in three scientific departments and 14 research groups as well as
common facilities for protein production and characterisation. The
department of matrix biology and tissue engineering consists of 6 research
groups, with interests ranging from structural biochemistry, cell biology,
development and tissue repair. Structural biology and bioinformatics are
also major areas of expertise. The IBCP is part of a grouping of local
institutes in the Gerland area, the IFR 128 Biosciences Lyon- Gerland
(www.ifr128.prd.fr). Lyon is one of the major cities in France
(www.lyon.fr/vdl/sections/ en) and is ideally located in the centre of
Europe within two hours of Paris, the Alps and the Mediterranean. It has a
strong cultural and intellectual tradition, is widely known as the
gastronomic centre of France, and the old town is a UNESCO World Heritage site.

Profile - Candidates should have a strong background in structural biology.
Knowledge of the French language would be helpful but not essential. There
are no restrictions on nationality.

[ccp4bb] How to improve the density of another molecule in Asymmetric unit

[Sun Tang]

Hello Everyone,
   
  I have a question about how to improve the electron density. I have two 
indepedent molecules in the asymmetric unit (1.9 A resolution). I solved the 
structure with PHASER and refined with CCP4i (Rfree = 0.29). 
   
  For the first molecule, the density is very good, but for the second one, the 
density is much worse than the first one. Are there any ways to improve the 
density of the second molecule, such as some kinds of averaging? Why that 
happens?
   
  Thank you very much for your help!
   
  Sincerely,
   
  Sun Tang
  [EMAIL PROTECTED] 

   
-
Never miss a thing.   Make Yahoo your homepage.

Re: [ccp4bb] crystallisation robot

[Lisa A Nagy]

You optimize that variable as well in scale-up.
That is the problem with nano-drop screening- sometimes it is near
impossible to scale up the drops. You have to re-screen around the
conditions (response surface methods can conserve experiments) because
the kinetics change so much between nano and big drops.

The smaller they are, the harder it is to scale up, so we compromise
between protein consumption and scalability. I think with most of these
machines, you _can_ tweak it to dispense in the 20-50nl range
accurately. To do that, you have to avoid using an air gap- either by
drawing (and diluting) your protein against a column of water, or by
using some other liquid (oil) between your protein and the dispensing
fluid. Many times these tiny drops yield crystals much faster because
they have more surface area to volume. But once you have them- well-
that's where I've pulled out my hair. Scale-up is hard, and for delicate
proteins (>1% crystalline hits), near impossible from the 20 nl drops.
That's the main reason we use bigger drops. 

In the early stages of a project, there is often a great deal of batch
to batch variability in the protein samples. Minimizing this, and
getting consistent polydispersity measurements can go a long way toward
making scale-ups easier.
All this screening is for naught if you need to optimize using a second
batch of protein which behaves differently from the first, or if your
first/only batch is degraded by the time you have hits to optimize. If
you are fortunate to be producing your own protein, you can personally
make sure you are producing consistent batches. The worst thing you can
do is screen on the hearts, then optimize/scaleup with the tails (of the
protein peak in the purification). 

Personally, I think having a liquid handling robot is almost as
important as a crystallization robot.

Lisa

P.S. Confidential to various sales people:
1. If you had visited us, you would have known we were going to buy
another robot. 
2. If you had visited us, you would have known I was in the group.
3. If a member of our group didn't contact you and request information
for purchasing your fabulous robot based on your website/mailings/ads,
then your marketing people (not us) have the problem.
4. Sole source justification. Features and Price. 20K is a significant
difference, and we didn't need the extra features on your machine. Or we
needed features yours didn't have. 
5. Sending snarky email messages about me to my coworkers makes all of
us less inclined to view your company favorably for future purchases. 

--
Lisa A. Nagy, Ph.D.
University of Alabama-Birmingham
[EMAIL PROTECTED]



-Original Message-
From: CCP4 bulletin board [mailto:[EMAIL PROTECTED] On Behalf Of
Oganesyan, Vaheh
Sent: Wednesday, January 16, 2008 9:06 AM
To: CCP4BB@JISCMAIL.AC.UK
Subject: Re: [ccp4bb] crystallisation robot
...
Once in a while I get large crystal that can be used for data
collection, but in most of the cases optimization in hanging 1+1 uL
drops is required.
What I found quite difficult to do is to choose the appropriate protein
concentration when moving from 200+200nL to 1+1 uL. Sometimes protein
should be diluted 3-4 times, sometimes it shouldn't. How others are
approaching this issue?

Vaheh

Re: [ccp4bb] crystallisation robot

[Shirley Roberts]

Vaheh,
If I'm optimising in 24 well trays after a hit in 96 I generally use 
sitting drop trays (cryschem) as a first step and try different ratios 
of protein:mother liquor first rather than changing the protein 
concentration. I know that sometimes the crystals stick to the plastic 
but I use the hypodermic trick mentioned on the BB a few days ago.


On the subject of drop size - I was at the RAMC meeting where this was 
discussed and several attendees had looked at the issue,  300+300nl or 
400+400nl were mentioned as more optimal drop sizes for obtaining 
crystal hits during screening.See summary on:

http://www.hamptonresearch.com/stuff/RAMC/RAMC2007Notes.aspx

At York we use 150+150nl  routinely for screening using the mosquito. 
I've moved up to 200+200 since the conference!


Shirley Roberts
ps I think I was responsible for the z to s change in the title
begin:vcard
fn:Shirley M.  Roberts
n:Roberts;Shirley M. 
org:York Structural Biology Laboratory
adr:University of York;;Chemistry Department;York;;YO10 5YW;UK
email;internet:[EMAIL PROTECTED]
tel;work:01904 328268
tel;fax:01904 328266
url:http://www.ysbl.york.ac.uk/people/5.htm
version:2.1
end:vcard

Re: [ccp4bb] crystallisation robot

[Oganesyan, Vaheh]

I have been using Phoenix for more than two years and so far there were
no issues with maintenance. Wash the needles and nano-dispenser before
and after the runs and you are good to go.
Electrostatic effects have been seen in a way of drops being positioned
on the side of the flat bottom plate, but the drops do not climb too
high in my case probably because I'm using fairly "large" drops (200 +
200 nl).
No matter how robust the system is, there will be always a way to break
it.

Once in a while I get large crystal that can be used for data
collection, but in most of the cases optimization in hanging 1+1 uL
drops is required.
What I found quite difficult to do is to choose the appropriate protein
concentration when moving from 200+200nL to 1+1 uL. Sometimes protein
should be diluted 3-4 times, sometimes it shouldn't. How others are
approaching this issue?

Vaheh

-Original Message-
From: CCP4 bulletin board [mailto:[EMAIL PROTECTED] On Behalf Of
Anastassis Perrakis
Sent: Wednesday, January 16, 2008 8:23 AM
To: CCP4BB@JISCMAIL.AC.UK
Subject: Re: [ccp4bb] crystallisation robot

> More recently, I've looked at all of the crystallization robot  
> vendors.
> For single lab users, all of the systems work well. Systems like  
> the Hydra
> or Mosquito are less automatic, but provide the basic functions for
> crystallization trial setup. For more of a user facility with a large
> number of users from several groups, you want more automation to avoid
> protocols that can damage the components, like alignment or  
> breakage of
> the needles. You also want to consider the total annual cost of
> expendables and maintenance.

There are two major kinds of  "a user facility with a large number of  
users from several groups"

a. One with an operator: then buy what the operator likes ;-)  
basically all machines do work
b. NO operator: buy something that is hard to break and needs minimal  
maintenance

We are in b. and happy with our Mosquito for 4 years or more now,  
although I am sure the
other choices mentioned are also clearly fine.

A.

Re: [ccp4bb] crystallisation robot - volume?

[cdekker]

Regarding stochastic processes: to increase the chances of nucleation 
one would like to have protein in the right nucleation zone for a 
certain period of time.


If the drop becomes too small, the ratio evaporation surface/drop 
volume changes. So, the nucleation zone might be reached within the 
drop but either too fast  ('racing through the phase diagram too 
quickly') or in the smaller droplet turbulence caused by evaporation 
might be more pronounced and interfere with nucleation.


At some stage I looked at this myself, although not for tiny volumes 
(0.3, 0.5 and 0.8ul droplets). With smaller volume I saw more and 
faster nucleation, but less crystal growth. There's got to be an 
optimum in terms of drop volume when using vapour diffusion. Any 
comments?


Carien

On 16 Jan 2008, at 13:17, Anastassis Perrakis wrote:


Oryxnano 50+50 nL

Demetres



Which, indirectly, brings up an interesting (but not relevant to the 
Oryx) question.


Nucleation is a process that does have a stochastic aspect.

Thus, one could argue that compromising to 200-300 nl might be better 
than either extremes of 50nl (too small volume and less chance for 
nucleation) or 1000 nl (too much sample).


any comments ? (let the flames begin).

A.

PS1
another interesting issue that has has been hardly touched in these 
emails is the real sample loss: left in wells and not easy to recover, 
lost because of contamination with system liquid, etc ...


PS2
I see lots of people with new robots. please do have a look at the 
www.BIOXHIT.org page and if you have a few minutes to assemble a table 
we will be happy to add your specs to our pages. it can be a nice 
resource and it has already enough things and already one response to 
my last email ;-) To make life easier to potential contributors we can 
provide an Excel sheet to fill up with your specs - just ask.


On Jan 16, 2008, at 12:46, Demetres D. Leonidas wrote:



David Briggs wrote:

I'll defend the honour of the phoenix... (again)

Bernhard Rupp 100+100 nl
Dave Briggs (and all users at Univ of Manchester, UK) 100+100nl
Others..

Only time we have ANY problems is when the nano dispensing tip gets 
clogged. Often a good wash whilst still on the machine will clear 
the blockage.


Dave




--

David C. Briggs PhD
Father & Crystallographer
http://www.dbriggs.talktalk.net 
AIM ID: dbassophile



--
Demetres D. Leonidas, Ph.D.
Structural Biology & Chemistry Group
Institute of Organic and Pharmaceutical Chemistry
The National Hellenic Research Foundation
48, Vassileos Constantinou Avenue
Athens 116 35, Greece
==
Tel. +30 210 7273841 (office)
+30 210 7273895 (lab) Fax. +30 210 7273831
E-mail: [EMAIL PROTECTED]
URL: http://athena.eie.gr
==



The Institute of Cancer Research: Royal Cancer Hospital, a charitable Company 
Limited by Guarantee, Registered in England under Company No. 534147 with its 
Registered Office at 123 Old Brompton Road, London SW7 3RP.

This e-mail message is confidential and for use by the addressee only.  If the 
message is received by anyone other than the addressee, please return the 
message to the sender by replying to it and then delete the message from your 
computer and network.

Re: [ccp4bb] crystallisation robot

[Anastassis Perrakis]

More recently, I've looked at all of the crystallization robot  
vendors.
For single lab users, all of the systems work well. Systems like  
the Hydra

or Mosquito are less automatic, but provide the basic functions for
crystallization trial setup. For more of a user facility with a large
number of users from several groups, you want more automation to avoid
protocols that can damage the components, like alignment or  
breakage of

the needles. You also want to consider the total annual cost of
expendables and maintenance.


There are two major kinds of  "a user facility with a large number of  
users from several groups"


a. One with an operator: then buy what the operator likes ;-)  
basically all machines do work
b. NO operator: buy something that is hard to break and needs minimal  
maintenance


We are in b. and happy with our Mosquito for 4 years or more now,  
although I am sure the

other choices mentioned are also clearly fine.

A.

Re: [ccp4bb] crystallisation robot

[Anastassis Perrakis]

Oryxnano 50+50 nL

Demetres



Which, indirectly, brings up an interesting (but not relevant to the  
Oryx) question.


Nucleation is a process that does have a stochastic aspect.

Thus, one could argue that compromising to 200-300 nl might be better  
than either extremes of 50nl (too small volume and less chance for  
nucleation) or 1000 nl (too much sample).


any comments ? (let the flames begin).

A.

PS1
another interesting issue that has has been hardly touched in these  
emails is the real sample loss: left in wells and not easy to  
recover, lost because of contamination with system liquid, etc ...


PS2
I see lots of people with new robots. please do have a look at the  
www.BIOXHIT.org page and if you have a few minutes to assemble a  
table we will be happy to add your specs to our pages. it can be a  
nice resource and it has already enough things and already one  
response to my last email ;-) To make life easier to potential  
contributors we can provide an Excel sheet to fill up with your specs  
- just ask.


On Jan 16, 2008, at 12:46, Demetres D. Leonidas wrote:



David Briggs wrote:

I'll defend the honour of the phoenix... (again)

Bernhard Rupp 100+100 nl
Dave Briggs (and all users at Univ of Manchester, UK) 100+100nl
Others..

Only time we have ANY problems is when the nano dispensing tip  
gets clogged. Often a good wash whilst still on the machine will  
clear the blockage.


Dave




--

David C. Briggs PhD
Father & Crystallographer
http://www.dbriggs.talktalk.net 
AIM ID: dbassophile



--
Demetres D. Leonidas, Ph.D.
Structural Biology & Chemistry Group
Institute of Organic and Pharmaceutical Chemistry
The National Hellenic Research Foundation
48, Vassileos Constantinou Avenue
Athens 116 35, Greece
==
Tel. +30 210 7273841 (office)
+30 210 7273895 (lab) Fax. +30 210 7273831
E-mail: [EMAIL PROTECTED]
URL: http://athena.eie.gr
==

Re: [ccp4bb] crystallisation robot

[Santarsiero, Bernard D.]

In our original work on a prototype, we used the Cartesian technology. We
were able to dispense 10nL+10nL drops with a large range of viscosities
without difficulty. The main issue was to wash the tips after use to
prevent clogging. That was with a system that could dispense 1nL droplets.
The present systems are sub-nL now. We didn't have a problem with
electrostatics.

More recently, I've looked at all of the crystallization robot vendors.
For single lab users, all of the systems work well. Systems like the Hydra
or Mosquito are less automatic, but provide the basic functions for
crystallization trial setup. For more of a user facility with a large
number of users from several groups, you want more automation to avoid
protocols that can damage the components, like alignment or breakage of
the needles. You also want to consider the total annual cost of
expendables and maintenance.

Bernie Santarsiero

On Wed, January 16, 2008 5:46 am, Demetres D. Leonidas wrote:
> Oryxnano 50+50 nL
>
> Demetres
>
>
> David Briggs wrote:
>> I'll defend the honour of the phoenix... (again)
>> Bernhard Rupp 100+100 nl
>> Dave Briggs (and all users at Univ of Manchester, UK) 100+100nl Others..
>> Only time we have ANY problems is when the nano dispensing tip gets
clogged. Often a good wash whilst still on the machine will clear the
blockage.
>> Dave
>> --
>> 
>> David C. Briggs PhD
>> Father & Crystallographer
>> http://www.dbriggs.talktalk.net  AIM
ID: dbassophile
>> 
>
> --
> Demetres D. Leonidas, Ph.D.
> Structural Biology & Chemistry Group
> Institute of Organic and Pharmaceutical Chemistry
> The National Hellenic Research Foundation
> 48, Vassileos Constantinou Avenue
> Athens 116 35, Greece
> ==
> Tel. +30 210 7273841 (office)
>  +30 210 7273895 (lab)
> Fax. +30 210 7273831
> E-mail: [EMAIL PROTECTED]
> URL: http://athena.eie.gr
> ==
>

Re: [ccp4bb] Perl

[Peter Keller]

Hi all,

On Wed, 16 Jan 2008, Ian Tickle wrote:


I'm rather surprised that is the complete explanation, for the simple
reason that Perl (or any program for that matter) already inherits all
the environment variables from the shell in which it is run, including
PATH, CINCL, CLIBD_MON etc which would be needed to run pdbset, refmac
etc.  In fact sourcing ccp4.setup from within Perl in the way suggested
will be very inefficient if you have several system calls since you
would have to do it in *every* system call that runs a CCP4 program,
i.e. the variable settings are not saved from one system call to the
next.


I don't believe that just doing

   system("source ccp4.setup");

in a Perl script will have any effect: if you check the return status of the 
above "system" function call you will find that it returns an error status. As 
described in the Perl documentation for "system", you can do this easily as 
follows:


   system("source env.test") == 0 or die "system call failed: $?";

This is because when "system" has only one argument, and that argument 
contains no shell metacharacters, that argument is split into words and passed 
to the system call execvp (do "man execvp" for details). execvp requires a 
executable file to run (either a binary, or a script of some type), and 
"source" is not an executable file. It is a builtin command of the shells csh, 
tcsh and bash. There is no executable file or script in /usr/bin or anywhere 
else on your $PATH in a standard Unix-family OS (unless you have put one there 
yourself).


You could force "system" to execute in a shell instead of using execvp by 
including a metacharacter in the argument like this:


   system("source ccp4.setup;");

or use an explicit shell invocation:

   system("/bin/bash -c 'source ccp4.setup'");

As long as ccp4.setup contains no errors, this function will return a 
successful status, but even that will have no effect on your script. The 
"system" function forks a child process to execute its argument. Your script 
will then wait for the child process to terminate before continuing. Modifying 
the environment of a child process will not have any effect on the environment 
of its parent. You could do something like:


   system("source ccp4.setup; pdbset ...");

as Ed suggested, although see below for a portability issue. As Ian points out 
this can get inefficient, and shouldn't be necessary anyway (unless you want 
to use different versions of CCP4 at different places in your script).



As Ed pointed out you didn't say what the *exact* error message was when
you tried it the first time (which is always a good idea when reporting
a problem!).  You said that the CCP4 programs work normally from the
command line which implies that the environment is correctly set up, so
the only conclusion I can come to is that you didn't source ccp4.setup
in the shell that you were running Perl in.  I always do exactly that


I think that Ian is correct here: whether your script works depends on whether 
CCP4 has been set up in the shell where you run your script.



(in my .login startup script so it's only run once when I login - I use
tcsh), then just as a check in all my Perl scripts the first line is
always:

if(!defined($ENV{"CCP4"})){
 die("You must set up the CCP4 environment.\n");
}

Also note that your solution won't work anyway if the user's default
shell is csh or tcsh because the system call in Perl uses sh by default,
whereas ccp4.setup will normally have been configured to be compatible
with csh or tcsh.


Not only this, but standard /bin/sh doesn't have a "source" command. It works 
on many Linux distributions because /bin/sh is a link to bash, or otherwise 
has some bash functionality. For portability, you should use the standard 
Bourne-shell "." command:


   system(". ccp4.setup; pdbset ");

Hope that this makes things clearer,
Peter.



HTH

-- Ian


-Original Message-
From: [EMAIL PROTECTED]
[mailto:[EMAIL PROTECTED] On Behalf Of
[EMAIL PROTECTED]
Sent: 15 January 2008 20:37
To: Ed Hoeffner
Cc: CCP4BB@JISCMAIL.AC.UK
Subject: Re: Perl

Hi,

thanks for all the help!

system ("source ccp4.setup")

does the trick, every time before a ccp4 script is called,
the environmental
parameters have to be set up this way.


Quoting Ed Hoeffner <[EMAIL PROTECTED]>:


Hi

You don't include any error messages, so it's very difficult to
troubleshoot.

However, as a wild guess, I'd suggest you ensure that you

execute the ccp4

setup file and then try your command. I don't know perl,

but the call might

look something like system("source ccp4.setup ; pdbset

..."). As I recall,

there is a ccp4 setup file that has to be loaded first,

though you may have

to find it and adjust that part of the command accordingly

so the file can

be located.

Ed





--





Disclaimer
This communication is confidential and may contain privileged information 
intended solely for the named addressee(s). It may not be used or disclosed 
except for th

Re: [ccp4bb] combine incomplete data sets

[David Briggs]

Hello Yadong,

This is what I would do in your position.

1) Integrate everything in MOSFLM
2) Enforce consistent indexing using POINTLESS (unless I am mistaken, there
are alternative origin(s) in p321)
http://www.ccp4.ac.uk/dist/html/alternate_origins.html
ftp://ftp.ccp4.ac.uk/ccp4/6.0.2/prerelease/pointless.html
3) Bundle ALL integrated datasets into one .mtz file (being such to make
sure all the batch number do not conflict)(POINTLESS may do this for you
now)
4) Push through SCALA/Truncate in one giant run - selecting your 'best'
dataset as a reference.

Scala should use all the reflections you give it, not just the overlapping
ones

The rejection of redundant reflections should only really be done if you
have a very good reason - redundancy (hopefully) adds to the quality of your
data

Scala/Truncate will give you masses of stats for individual datasets, and
how well they scale together.

At the end you should get a single .mtz file containing all your reflections
scaled together and converted to Fs.

Hope this helps,

David


-- 

David C. Briggs PhD
Father & Crystallographer
http://www.dbriggs.talktalk.net
AIM ID: dbassophile

[ccp4bb] Job offer: Protein Crystallographer (with Ph.D)

[Blaesse, Michael]

PROTEROS is a leading service-provider for the preparation and
three-dimensional structural analysis of proteins for structure-guided
drug discovery. Our work contributes to fast and rational development of
new therapeutic compounds. Since its foundation in 1998, PROTEROS is a
continuously growing, renowned enterprise collaborating with a large
variety of international Pharma, Biotech and Crop Science companies.

To support our existing crystallography team we are looking for a

Protein Crystallographer (female/male)

 

Job description

You will be responsible for

- planning and execution of protein crystallization experiments

- accomplishment of small molecule ligand binding to protein crystals

- analysis and evaluation of protein crystallization experiments,
protein crystals, and protein X-ray diffraction patterns

- evaluation of protein X-ray diffraction data

- X-ray diffraction data collection at in-house generators and
synchrotron beamlines

- further development and optimization of methods and work-flows in
protein crystallization and protein crystal structure determination

- protein structure solution, modelling and refinement

 

The successful candidate will meet the following requirements

You have a Ph.D. in Chemistry, Biology, or Pharmacy with excellent
records providing you with a profound knowledge and experience in
protein crystallization, protein-ligand crystal complex formation and
protein structure determination. Knowledge and experience in
Fragment-based crystallography would be a plus. A good knowledge with
personal computers including UNIX/LINUX systems and acquaintance with
current software packages employed in protein crystallography are
mandatory.

You are proactive and able to work effectively under strict time lines.

Additionally, you have excellent communication skills and like to work
in a multi-disciplinary team. Industry experience in the life science
industry is an asset.

 

Please apply by sending your full application documents to

[EMAIL PROTECTED]

or to

Rabea Neumann | Am Klopferspitz 19 | 82152 Martinsried | Germany

 

[ccp4bb] combine incomplete data sets

[Yu, Yadong]

Hi there,

I have 3 incomplete data sets, collected from 3 single crystals, each accounts 
for ~40% completeness. They have same resolution range, and randomly overlap 
with each other a small portion.

Mosflm and Scala work well for all 3 separately. The 2 from same beamline have 
very close statistics, while the 3rd from another beamline differs drastically. 
Reflections occuring both in the 1st and 3rd show amplitudes with large 
variance in either direction. The truncate process could be inaccurate because 
of low resolution. Knowing this I still have a few questions: 

1. is there alternate origin of unit cell for space group p321? 
2. does scala use all reflections in the reference data set to scale another? 
or just those occuring both in 2?
3. when scaling a data with a reference, does program work out an overall scale 
factor compared to reference, or a series of scales for different resolution 
shells? 
4. when combining 2 data, do people simply throw away redundancies, or use them 
for averaging? 
5. once all 3 combined, is it possible to make a statistics for this combined 
data set? 

Any suggestions are highly appreciated.

Yadong

Yadong Yu
Postdoctoral scholar
Department of Biochemistry and Biophysics, UCSF
Genentech Hall, Rm316
600 16th ST,
San Francisco, CA 94158
415-514-9708
[EMAIL PROTECTED]

Re: [ccp4bb] crystallisation robot

[Demetres D. Leonidas]

Oryxnano 50+50 nL

Demetres


David Briggs wrote:

I'll defend the honour of the phoenix... (again)

Bernhard Rupp 100+100 nl
Dave Briggs (and all users at Univ of Manchester, UK) 100+100nl   


Others..

Only time we have ANY problems is when the nano dispensing tip gets 
clogged. Often a good wash whilst still on the machine will clear the 
blockage.


Dave




--

David C. Briggs PhD
Father & Crystallographer
http://www.dbriggs.talktalk.net 
AIM ID: dbassophile
 


--
Demetres D. Leonidas, Ph.D.
Structural Biology & Chemistry Group
Institute of Organic and Pharmaceutical Chemistry
The National Hellenic Research Foundation
48, Vassileos Constantinou Avenue
Athens 116 35, Greece
==
Tel. +30 210 7273841 (office)
+30 210 7273895 (lab) 
Fax. +30 210 7273831

E-mail: [EMAIL PROTECTED]
URL: http://athena.eie.gr
==

Re: [ccp4bb] crystallisation robot

[David Briggs]

I'll defend the honour of the phoenix... (again)

Bernhard Rupp 100+100 nl
Dave Briggs (and all users at Univ of Manchester, UK) 100+100nl

Others..
Only time we have ANY problems is when the nano dispensing tip gets clogged.
Often a good wash whilst still on the machine will clear the blockage.

Dave




-- 

David C. Briggs PhD
Father & Crystallographer
http://www.dbriggs.talktalk.net
AIM ID: dbassophile

Re: [ccp4bb] crystallisation robot

[Alexey Rak]

Dear Bernard,

I do not have long experience using Phoenix. I tried the machine during a 
couple of demonstrations I had chance to participate and also learned about 
other people experience who used the robot (various MPIs and universities in 
Germany and a couple of users from industry in Europe), I hope they can comment 
further. 

The crash of Phoenix needles that dissolved a myth on non-breakable Phoenix 
needles we witnessed during Art Robbins instruments presentation in PSDI 
meeting, 28th - 30th October 2007 Autrans, France. The needles were bended by 
rep during demonstration in the first day of the conference so that the robot 
head was moving without needles for the rest of the conference, it was funny.

For the maintenance, all labs I know those using Phoenix have someone looking 
after the machine. In some lab setups the situation exaggerated even further: 
lab members do not have direct access to the delicate machine and all the 
crystallization by Phoenix is carried out by person who is trained as the User 
and dedicated to. As being quite experienced user of Cartesian nano dispenser 
(1 year), I can understand the problems of cleanness for the robots with not 
dispensable and therefore requiring washing steps needles/tips. As I know, the 
situation is significantly improved in Innovadyne screenmaker, for those who 
prefer flexibilities of 96 channel system over speed, simplicity, no cross 
contamination, accuracy of pipetting high PEGs Mosquito system (4 years 
experience).

The electrostatic effect I mentioned was observed on the plates performed by 
Phoenix, while setting the same plates (Greiner triple, Corning 3550) by 
Mosquito did not cause similar problems (same drops volumes, same screening 
solutions), I hope colleagues may detail further. I would also suggest 
experienced Phoenix users to let us know what drops volume is routinely set in 
their labs.

Bernhard Rupp 100+100 nl
Others??? 
Others..

All the best,
Alexey.

 

-Original Message-
From: CCP4 bulletin board [mailto:[EMAIL PROTECTED] On Behalf Of Bernhard Rupp
Sent: Wednesday, January 16, 2008 1:53 AM
To: CCP4BB@JISCMAIL.AC.UK
Subject: Re: [ccp4bb] crystallisation robot

Dear Alexey,

being involved in the development of the 'fixed needles + a few' 
robots and 96-well plates early on, I wonder about your bad experiences. 

You seem to say that the Phoenix requires more maintenance than the Mosquito - 
how long have you had that Phoenix or what model is giving you the trouble?

For the electrostatics issue: it’s the plates that are electrostatic not the 
robot. This is a common lament for plastic plate users, and the Intelliplate 
designed to go with the Phoenix comes in anti-static bags as used in electronic 
component packaging. Other plates have anti-static coating, and there may be 
other tricks as well to avoid electrostatic charge 
(comments anyone?)
  
Also, the limitation of the 200+200 you claim for the Phoenix seems extreme, 
users have set up 200+200 already with the old hydra+1 contraptions, and 
100+100 seems to work reliable with the Phoenix (maybe some users can 
comment?). Could it be that yours is not set up correctly? I am sure ARI will 
be happy to send a European rep by to check.

Best regards, BR 

-Original Message-
From: CCP4 bulletin board [mailto:[EMAIL PROTECTED] On Behalf Of Alexey Rak
Sent: Monday, January 14, 2008 9:36 AM
To: CCP4BB@JISCMAIL.AC.UK
Subject: Re: [ccp4bb] crystallisation robot

we have recently tried the new lid mentioned below. It works very well, 
Mosquito makes 3 hanging drops 96 well plate in 3-4 minutes! The film is very 
easy to handle and it is very transparent. The price for the lid is 
significantly reduced: from 13€ before to 4€ now.

Comparing to Cartesian and Phoenix robots the Mosquito is REALLY maintenance 
free machine, especially the third generation insect where many 1st and 2nd 
Mosquito generation users advices have been implicated.

Alexey Rak
Structural Biology, Chemical Sciences
Sanofi-Aventis
Centre de Recherche Paris

Re: [ccp4bb] Perl

[Ian Tickle]

I'm rather surprised that is the complete explanation, for the simple
reason that Perl (or any program for that matter) already inherits all
the environment variables from the shell in which it is run, including
PATH, CINCL, CLIBD_MON etc which would be needed to run pdbset, refmac
etc.  In fact sourcing ccp4.setup from within Perl in the way suggested
will be very inefficient if you have several system calls since you
would have to do it in *every* system call that runs a CCP4 program,
i.e. the variable settings are not saved from one system call to the
next.

As Ed pointed out you didn't say what the *exact* error message was when
you tried it the first time (which is always a good idea when reporting
a problem!).  You said that the CCP4 programs work normally from the
command line which implies that the environment is correctly set up, so
the only conclusion I can come to is that you didn't source ccp4.setup
in the shell that you were running Perl in.  I always do exactly that
(in my .login startup script so it's only run once when I login - I use
tcsh), then just as a check in all my Perl scripts the first line is
always:

if(!defined($ENV{"CCP4"})){
  die("You must set up the CCP4 environment.\n");
}

Also note that your solution won't work anyway if the user's default
shell is csh or tcsh because the system call in Perl uses sh by default,
whereas ccp4.setup will normally have been configured to be compatible
with csh or tcsh.

HTH

-- Ian

> -Original Message-
> From: [EMAIL PROTECTED] 
> [mailto:[EMAIL PROTECTED] On Behalf Of 
> [EMAIL PROTECTED]
> Sent: 15 January 2008 20:37
> To: Ed Hoeffner
> Cc: CCP4BB@JISCMAIL.AC.UK
> Subject: Re: Perl
> 
> Hi,
> 
> thanks for all the help!
> 
> system ("source ccp4.setup")
> 
> does the trick, every time before a ccp4 script is called, 
> the environmental
> parameters have to be set up this way.
> 
> 
> Quoting Ed Hoeffner <[EMAIL PROTECTED]>:
> 
> > Hi
> > 
> > You don't include any error messages, so it's very difficult to
> > troubleshoot.
> > 
> > However, as a wild guess, I'd suggest you ensure that you 
> execute the ccp4
> > setup file and then try your command. I don't know perl, 
> but the call might
> > look something like system("source ccp4.setup ; pdbset 
> ..."). As I recall,
> > there is a ccp4 setup file that has to be loaded first, 
> though you may have
> > to find it and adjust that part of the command accordingly 
> so the file can
> > be located.
> > 
> > Ed
> > 
> > 
> 
> 
> -- 
> 
> 


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Re: [ccp4bb] Calculating volume of Ligands

[Isaac Westwood]

 

Dear Rajan,

 

In addition to the estimates already mentioned, if you have access to Chem3D
(Cambridgesoft), you can calculate the Connolly volume (excluding solvent)
for any (preferably small) molecule, either drawn by hand or by input from
its coordinates. In ChemBio3D ultra, this is found in calculations >
computer properties > ChemPropStd.

Another guestimate method is to determine the distances between various
pairs of atoms in, e.g. Coot, and to do a back-of-an-envelope calculation
for the volume based on those, but that's probably less accurate than the
estimate methods already mentioned!

 

Isaac

 

--

Dr Isaac Westwood

Department of Pharmacology

University of Oxford

Mansfield Road

Oxford

OX1 3QT

tel: 01865 271595

email: [EMAIL PROTECTED]

 

  _  

From: CCP4 bulletin board [mailto:[EMAIL PROTECTED] On Behalf Of Rajan
Pillai
Sent: 15 January 2008 21:40
To: CCP4BB@JISCMAIL.AC.UK
Subject: [ccp4bb] Calculating volume of Ligands

 

Hi All,

Can anyone tell me any program that calculates voume of a ligand? Moreover,
is there also any program that can calculate the volume of a ligand from its
coordinates?

Thanks,

Rajan.

Re: [ccp4bb] Calculating volume of Ligands

[Eleanor Dodson]

It isnt a bad approximation for any organic compound to take the number 
of atoms including hydrogens, and multiply it by 10A^^3

Eleanor Dodson

Rajan Pillai wrote:

Hi All,

Can anyone tell me any program that calculates voume of a ligand? Moreover,
is there also any program that can calculate the volume of a ligand from its
coordinates?

Thanks,

Rajan.

  

[ccp4bb] lsqkab

[Nathalie Colloc'h]

Hi everybody,

I wonder why in lsqkab, the glycine residues have a rms value for their 
side chains which are non null
(the same calculation done with CNS give about the same results for the 
rms calculation, but with

null value for the rms of glycine side chains).

thanks a lot

nathalie

--
Dr. Nathalie Colloc'h, PhD
CI-NAPS, UMR 6232 - UCBN - CNRS
GIP Cyceron
Bd Becquerel, BP5229
14074 Caen cedex
FRANCE
Tel. 33.2.31.47.01.32
Fax. 33.2.31.47.02.22
[EMAIL PROTECTED]

Re: [ccp4bb] Calculating volume of Ligands

[harry powell]

Hi

A simple trick that small molecule crystallographers have been using  
for decades is based on the average volume of non-hydrogen atoms  
being about 18 Å^3 (this is close to being more-or-less correct for  
C, N and O, and the presence of one or two S or P atoms usually makes  
little difference) - they simply multiply the number of non-H atoms  
in the ligand by 18 and - hey presto!


If you've only got a few atoms in your ligand, this is dead easy and  
you don't need a program to do it. If you're in a hurry, use 20Å^3  
instead for a slight overestimate...


On 15 Jan 2008, at 21:39, Rajan Pillai wrote:


Hi All,

Can anyone tell me any program that calculates voume of a ligand?  
Moreover, is there also any program that can calculate the volume  
of a ligand from its coordinates?


Thanks,

Rajan.


Harry
--
Dr Harry Powell, MRC Laboratory of Molecular Biology, MRC Centre, Hills
Road, Cambridge, CB2 2QH