Hi Jacob,
take some (few ul) of the lysed culture and spread that together with regular
cells onto an agar plate so that you (would) get a lawn of bacteria after O/N
incubation. If it is phage you'll get clear plaques in your lawn, very
distinctive. If it is regular cell lysis induced by the to
Yes,
This is 95% likely to be the dreaded T1 phage (well, OK - any lytic phage
that infects your culture, really). The almost-only other alternative is
that your protein lyses the cells.
Dealing with phage infection can be tough especially in the multi-user
environment and doubly so if you make y
Dear Jacob,
In my hands, different strains of E. coli appear to behave differently
- I found BL21(DE3) needing harder centrifugation than JM109(DE3) to
get a good pellet. And BL21 lysing partially when doing osmotic shock
extraction of a periplasmically expressed protein - while XL1Blue
be
Okay, it seems that the consensus is phage infection. Is there anything to
seal the diagnosis? Also, does anybody have literature on de-phaging
glassware? I am assuming that regular autoclaving will not do the trick?
Jacob
***
Jacob Pearson Keller
Northw
The same is true for Porphyridium purpureum
beta-CA and Halothiobacillus neapolitanus beta-CA. Both are composed of
pseudodimers composed of two structurally homologous domains. In the
case of H. neapolitanus, the domains have very little sequence
homology, and one domain has lost its active si
Hi all,
another reason to get low readings from the UV meter can be a drift
in the baseline. The photometer is properly calibrated only when
you turn it on, I think. Since we have our machine running in the
coldroom without (usually) turning it off for month, we have observed
wrong 260/280 r
Dear Peter,
it's a common phenomenon to create protein concentration gradients inside a
protein concentrator. Simply take it all out of the concentrator, put it in
a separate tube, and mix thoroughly/vortex. The 'slime' may very well
redissolve and you'll a homogeneous distribution. What you des
Dear Jacob,
these are the hallmark signs for a bacteriophage infection. I'm afraid
you'll have lots of bleaching / baking of glassware to do...
Once you have them in the lab, they're very hard to get rid of. You can
test one of your previous constructs that didn't lyse the cells; if these
now d
Dear crystallographers,
I recently expressed some new constructs, and found after my usual
expression protocol that the cell pellets were not compacted at the bottom
corner of the bottles us usual, but were instead smeared as a film on the
side, and further, were somewhat clumpy, like clots, a
Peter,
If I understand what you are saying - then it is very likely that your
protein forms aggregates. Whether this happens on concentration or not is
unknown because concentration may simply bring the pre-existing aggregates
to the membrane.
You can try to make concentration process 'easy
Identical is a very strong term - however if you're looking for 'very
similar' instead of identical then you should look at any kind of
extracellular receptor with Ig-like or Efhand-like domains - these often
look like choo-choo trains. Receptor-like phosphatases have similar
features. Another frui
Hello all
I am working with a small protein-protein complex. This complex express
quite well . I purify in a buffer of pH=9.0 with 150mM NaCl and 1% of
glycerol and able to concentrate upto 20 mg per ml. I have a two clones of
this protein complex. One is N-terminal His tagged and another C-termin
The Gordon Research Conference on Diffraction Methods in Structural
Biology will be held at Bates College in Lewiston, Maine, from July
18-23, 2010.
As in the past, the program will include the latest developments in
methodology covering all aspects of macromolecular crystallography,
from cry
The cadmium-utilizing marine diatom carbonic anhydrase (CA) protein has
three consecutive CA domains that have very similar structures but
non-identical sequences.
See:
Structure and metal exchange in the cadmium carbonic anhydrase of marine
diatoms.
Xu Y, Feng L, Jeffrey PD, Shi Y, Morel FM.
Dear all,
I would like to draw your attention to the following job announcement:
Position
Research technician for protein expression
Closing date: 02.08.2009
On the web:
http://www.embl-hamburg.de/aboutus/jobs/jobs_embl_hamburg/2009/w_09_050_RT_Protein/index.html
Job Description
The EMBL
Hi Shankar,
Another fascinating example might be Dscam (Down Syndrome cell adhesion
molecule) with multiple Ig like domains and Fibronectin type III domains.
Different splice isoforms are expressed in different nerves, and it serves
as a recognition module, same repels, different ones attract.
No
Hi Shankar
gamma-crystallin is likely a good candidate [Blundell et al. Nature
289:771-777, 1981], and it in fact goes a step further. It combines two
nearly identical greek-key motifs in each domain, which are then connected
via a loop to make a dimer. This is all in one chain.
Best wishes
Sa
*Post-doctoral position within structural studies on cytoskeletal
proteins at the Division of Structural Biology at the Helmholtz Centre
for Infection in t**he research group of Asst. Prof. Inari Kursula at
the University of Hamburg/DESY, Hamburg, Germany*
The successful candidate, possessing
thank you all for quick answers.
-shankar
On Thu, Jul 2, 2009 at 12:47 PM, Charlie Bond wrote:
> Depending on your definition of 'identical', examples of repeated gene
> duplication contribute to these:
>
> You could look at glyoxalases where there are dimeric examples where each
> monomer is co
Depending on your definition of 'identical', examples of repeated gene
duplication contribute to these:
You could look at glyoxalases where there are dimeric examples where
each monomer is composed of a repeated subdomain (A1-A2:A1-A2) and
monomeric examples where a further duplication has occ
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