Nothing out of the ordinary there.
You can try to induce overnight (for 20 hours or so) at 15C. And you can
try to reduce the concentration of IPTG if excess protein is going to
inclusion bodies anyways. Collect whatever protein is soluble and ignore
your inclusion bodies. The His tags allow you
Ian Tickle replied:
(freezing soaking can easily induce 1% and sometimes 5% change),
is surely the reason that Fo-Fo maps never caught on!
I've been curious about this unit cell constant mis-matching for a
while. If I understood well, Perutz tried to exploit the effect
Dear all,
i have protein sequence and how can i predict the secondary structure from the
sequence..???
Thank u
hsn.
Hi Hussain,
Psipred is a good a place as any to start:
http://bioinf.cs.ucl.ac.uk/psipred/
Cheers,
Dave
David C. Briggs PhD
Father, Structural Biologist and Sceptic
University of Manchester E-mail:
david.c.bri...@manchester.ac.uk
Exapsy (http://www.expasy.ch/tools/#proteome) also has a whole section about
secondary structure prediction.
Tim
On Wed, May 05, 2010 at 04:20:13PM +0530, Hussain Bhukyagps wrote:
Dear all,
i have protein sequence and how can i predict the secondary structure from
the sequence..???
Hi -
I am refining a low (3.7Å) structure with refmac(5.5.0091), and am
having trouble maintaining some secondary structure elements. I would
like to restrain the H-bonding in clear secondary structural
elements, which should help prevent carbonyls in helices from
flipping out, etc, but
Several suggestions:
Try tagging with a solubility-enhancing tag
like GST or NusA.
If using a pET vector system, try "leaky"
_expression_. The T7 promoter is not well-repressed (except in pLysS
systems) and you can get low but very significant levels of _expression_
without induction.
try JM109(DE3) instead of BL21(DE3), combined with low temperature induction -
for several proteins this has yielded us soluble protein instead of inclusion
bodies. I think this is because JM109(DE3) grows a bit slower.
Mark J van Raaij
mvr...@ibmb.csic.es
Hi Greg,
you can generate secondary structure restraints definitions for Refmac
using this command:
phenix.secondary_structure_restraints model.pdb format=refmac
Pavel.
On 5/5/10 4:33 AM, Gregory Bowman wrote:
Hi -
I am refining a low (3.7Å) structure with refmac(5.5.0091), and am
Hi,
I have problems solving the structure of a protein crystal which seems to be
disordered. In order to investigate the disorder it would be useful to have a
precision photograph that shows reflections only in the [0kl] plane. Does
anyone know software that can transform raw data to give
hklview will generate pseudo precession images
Sent from my iPhone
On 05/05/2010, at 11:03 PM, Tillmann Heinisch tillmann.heini...@unibas.ch
wrote:
Hi,
I have problems solving the structure of a protein crystal which
seems to be disordered. In order to investigate the disorder it
would
to my knowledge hklview just works with integrated data whereas I need to plot
raw intensities along h, k and l to investigate reflection streakings. I heard
such software is routinely used in small molecule crystallography.
Tillmann
On May 5, 2010, at 3:18 PM, David Briggs wrote:
Hi
I have found that if you give a cold shock ( 4 degrees for 30 mins-1 hr)
before low temperature induction it helps to keep proteins
soluble.
Ivan
Hi Tillmann,
what do you mean by 'raw intensities' as opposed to integrated data?
Would xprep be an option for you? It reads XDS_ASCII.HKL, but that's of course
after integration.
But it should be easy to convert any (non-binary) file containing raw
intensities into an hkl-file that you can read
I think he's looking for a program which will extract a plane from the raw 3D
reciprocal space, as sampled by the raw images (ie before integration, but with
the plane defined by the indexed lattice). That's a much harder job
Phil
On 5 May 2010, at 16:50, Tim Gruene wrote:
Hi Tillmann,
what
As Phil says, constructing an undistorted slice through reciprocal space
is much harder than displaying integrated intensities. The Bruker APEX2
software does this nicely and I understand that they can also convert
MAR CCD and possibly some other frame formats to Bruker format, which
presumably
Hi
this is what I thought. I would imagine that you'd want to combine the
2D information in the X-ray images in a dataset into a 3D solid
figure, then look at slices through this for the RL projection you
were interested in. If you indexed the dataset first you should be
able to get the
This 'much harder job' is accomplished, too, in the comprehensive
CrysAlisPro
data-analysis suite from Oxford Diffraction. Again: not just from
images in
the OD format, but from a wide variety of others, also. So, if we could
assist in
this case, Tillmann, then we would be pleased
Storing a complete 3D image set in memory is moderately challenging even for
today's computers (probably several to several 10s of gigabytes, though you
could perhaps cheat a bit), so programs would probably have to use
old-fashioned double sort techniques to extract a zone. I wonder how the
I wrote a little jiffy for doing this some years ago:
http://bl831.als.lbl.gov/~jamesh/pickup/adsc2pdb.com
However, I should note that this program relies on the DPS program:
dps_peaksearch to pick spots on each image in your data set. These
spots are then transformed into reciprocal space
On Wednesday 05 May 2010 12:49:34 am Jon Wright wrote:
I've been curious about this unit cell constant mis-matching for a
while. If I understood well, Perutz tried to exploit the effect for
phasing prior to heavy atom methods. As the unit cell changes, the
diffraction peaks move,
English et al. Recombinant and in vitro expression systems for hydrogenases:
new frontiers in basic and applied studies for biological and synthetic H2
production. Dalton Trans. (2009) (45) pp. 9970-8
I've never tried to express a hydrogenase, but I've read that it's very
hard. Do you have all
Tillmann,
I wrote a little jiffy some months ago, to go from raw images to a
pseudo-precession photo, in the context of labelit.index. I'll dig it
out and post a link to the program...
Nick
Tillmann Heinisch wrote:
Hi,
I have problems solving the structure of a protein crystal which
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