[ccp4bb] Postdoctoral Position of Membrane Protein Structural Biology

[Hu, Jian]

Description:

A postdoctoral research associate position in membrane protein structural 
biology/biochemistry is immediately available in the Department of Biochemistry 
& Molecular Biology at Michigan State University, East Lansing, MI, US. One 
major direction in Dr. Jian Hu’s laboratory is structural biology study of 
metal transporters important in medicine. Previously, we have solved the first 
crystal structure of the extracellular domain of a mammalian ZIP4 zinc 
transporter (Nature Communications, 2016) and the first full length ZIP metal 
transporter (Science Advances, 2017). By employing multidisciplinary approaches 
of structural biology, biochemistry, biophysics and cell biology, we are aiming 
to establish the structural basis and clarify the molecular mechanisms of metal 
homeostasis in eukaryotic cells. The MSU cryo-EM facility will be updated with 
a new transmission and scanning transmission electron microscope by the end of 
2018. The laboratory is well funded by federal agencies, and the position can 
be supported up to four years based on mutual satisfaction. The other ongoing 
projects include biogenesis of the Ni-pincer cofactor, the PIPK lipid kinases 
and the calcium sensing receptor. To obtain more information, please visit our 
website (https://bmb.natsci.msu.edu/faculty/jian-hu/).

The successful candidate must be highly self-motivated with Ph.D. in structural 
biology, biochemistry, biophysics, molecular biology or related fields. 
Hands-on experience in membrane protein biochemistry is required. Experience in 
structural biology (x-ray crystallography/cryo-EM/NMR) is desirable, but not 
necessary. Interested individuals should submit a brief cover letter, CV, and 
the names and the contact information of three referees to Dr. Jian Hu by 
email: huji...@msu.edu.


~~

Jian Hu, PhD
Assistant Professor,
Department of Biochemistry and Molecular Biology &
Department of Chemistry,
Michigan State University,
603 Wilson Rd., East Lansing, MI 48824
https://bmb.natsci.msu.edu/faculty/jian-hu/



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Re: [ccp4bb] Normalization of B-factors

[Ethan Merritt]

On Thursday, 09 August 2018 10:45:07 Pavel Afonine wrote:
> > I (personally) think the best answer from these was to look at the
> > TLS-subtracted residuals (ie. total B-factor - TLS component) — can’t
> > remember who sent it, off the top of my head.
> >
> 
> TLS is just an approximation, sometimes good and sometimes not. If TLS
> parameters are refined along with individual ADPs ("residual") the latter
> tend to compensate for eventual inadequacy of TLS model.
> 
> Pavel

Depending on the quality and resolution of your data, a different summary
may apply

  If individual ADPs are refined along with a TLS model they may add
  noise that obscures the significance of bulk motion and may 
  artifactually reduce R-factors through over-fitting. 


The best method of comparing structures ultimately depends on what
question you are trying to answer.

If you are trying to document a reduction in overall flexibility due to
ligand binding, the TLS descriptions (rather than individual B factors)
may be the most informative thing to compare.  If you are looking
for specific residues that become "locked down" or "disengaged"
upon ligand binding, then the opposite is true: you would want to
compare the residual B-factors in those residues after subtracting
out the TLS contribution.

At sufficiently high resolution you should also look for evidence
that some residues may gain or lose alternate conformations upon
ligand binding.  High B factors may indicate that an alternate
conformation has been missed, or its occupancy may have changed.

Ethan

-- 
Ethan A Merritt, Dept of Biochemistry
Biomolecular Structure Center,  K-428 Health Sciences Bldg
MS 357742,   University of Washington, Seattle 98195-7742



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Re: [ccp4bb] Normalization of B-factors

[Eleanor Dodson]

I wouldnt have thought so.
There is a B factor plot as part of CCP4I2 - does that show any pattern of
differences between ligand and environ,ment?

Eleanor

On 9 August 2018 at 10:54, Santhosh Gatreddy 
wrote:

> Hi all,
>
> I have to compare the B-factors of three of my ligand bound structures of
> the same protein which were diffracted to 1.85, 1.98 and 2.01 A resolution.
>
> Is it necessary to normalize the B-factors of these structures (dimer in
> ASU) before comparing them to understand the ligand induced stability.
>
>
> Thanks for Your suggestions,
>
> Santhosh
>
>
>
>
>
>
>
>
>
> 
>
> To unsubscribe from the CCP4BB list, click the following link:
> https://www.jiscmail.ac.uk/cgi-bin/webadmin?SUBED1=CCP4BB=1
>



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Re: [ccp4bb] screw axes /system. absenses and phaser/MR solutions

[Eleanor Dodson]

Well that is pretty obviously P 2 21 2 - h 0 0   and l 0 0 are obviously
present..

The 0 k 0 absences could be generated by a nc translation of x, 1/2, z - do
you have that?

Twinning is unusual in P2/mmm  but possible of course - can you send the
pointless log file?
Eleanor



On 9 August 2018 at 09:11, Kajander, Tommi A 
wrote:

> Hi Tim - There are reflections in all directions and only one clearly has
> systematic absenses. Although only few seem to be present in one direction
> (But thats the h00 direction here
> + all are present though) -phaser finds with same data P22121 as the
> strongest solution (few reflections are weak in that direction from below…
> but most not).  TZ-score diffrence is not
> hure 13 vs 15.
>
> P 2 2 21
>   (   0,0,5): i/sigi =   20.8
>   (   0,0,7): i/sigi =   19.6
>   (   0,0,9): i/sigi =   21.0
>   (   0,0,   11): i/sigi =1.9
>   (   0,0,   13): i/sigi =   21.0
>   (   0,0,   15): i/sigi =6.0
>   (   0,0,   17): i/sigi =   20.8
>   (   0,0,   19): i/sigi =0.9
>   (   0,0,   21): i/sigi =9.0
>   (   0,0,   23): i/sigi =1.0
>   (   0,0,   25): i/sigi =4.9
>   (   0,0,   27): i/sigi =   19.9
>   (   0,0,   29): i/sigi =   20.9
>   (   0,0,   31): i/sigi =   20.2
>   (   0,0,   33): i/sigi =   19.1
>   (   0,0,   35): i/sigi =   18.2
>   (   0,0,   37): i/sigi =   13.1
>   (   0,0,   39): i/sigi =   20.8
>   (   0,0,   41): i/sigi =8.9
>   (   0,0,   43): i/sigi =   12.7
>   (   0,0,   45): i/sigi =2.5
>   (   0,0,   47): i/sigi =0.8
> P 21 2 2
>   (   3,0,0): i/sigi =   31.2
>   (   5,0,0): i/sigi =   25.2
>   (   7,0,0): i/sigi =   21.0
>   (   9,0,0): i/sigi =2.6
> P 2 21 2
>   (   0,5,0): i/sigi =0.7
>   (   0,7,0): i/sigi =0.7
>   (   0,9,0): i/sigi =0.7
>   (   0,   11,0): i/sigi =1.1
>   (   0,   13,0): i/sigi =1.0
>   (   0,   15,0): i/sigi =1.5
>   (   0,   17,0): i/sigi =0.7
>   (   0,   19,0): i/sigi =0.7
>   (   0,   21,0): i/sigi =0.7
>   (   0,   23,0): i/sigi =0.8
>   (   0,   25,0): i/sigi =0.7
>   (   0,   29,0): i/sigi =0.7
>   (   0,   31,0): i/sigi =0.8
>   (   0,   33,0): i/sigi =0.7
>   (   0,   35,0): i/sigi =0.7
>   (   0,   37,0): i/sigi =0.9
>   (   0,   39,0): i/sigi =0.7
>   (   0,   41,0): i/sigi =0.7
>   (   0,   43,0): i/sigi =0.7
>
>
> Neither seem to refine now so have to work on it a bit more beyond this  -
> we see the same “conflict" with two different crystals - seems complicated
> - the other one processed much
> better in P21  and seemed possibly twinned. Maybe the case here too.
>
> Tommi
>
>
>
>
>
> On Aug 9, 2018, at 10:29 AM, Tim Gruene  wrote:
>
> Dear Tommi,
>
> did you check whether you collected any reflections at all that should
> be absent for the second screw axis? If there are non - which could
> easily happen with low resolution, incomplete data - pointless and XDS
> might be conservative and not estimate the likelihood for the second
> screw-axis.
>
> MR, however, will pick up the space group with all data, and thus be
> able to tell between P2212 and P22121 whether or not you recorded
> reflections that are expected to be absent.
>
> Best,
> Tim
>
> On 08/08/2018 04:29 PM, Kajander, Tommi A wrote:
>
> Hi,
> Any clues why the followting happens: pointless (and just looking at the
> XDS output) clearly tells there is one screw axis in P-ortorhombic (P2212)
> yet phaser gives the best Z-scores in P22121. (...I suspect this may be to
> do with twinning - might be monoclinic twiined still though now processes
> very well in P222.)
>
> If i run the mtz after XDSCONV (ie F2MTZ) via pointless (instead of
> directly after XDS) it also suggests this - but i suppose i am not suppose
> to run merged data via pointless.
>
> Thanks for comments,
>
> Tommi
>
> 
>
> To unsubscribe from the CCP4BB list, click the following link:
> https://www.jiscmail.ac.uk/cgi-bin/webadmin?SUBED1=CCP4BB=1
>
>
> --
> --
> Paul Scherrer Institut
> Tim Gruene
> - persoenlich -
> OSUA/204
> CH-5232 Villigen PSI
> phone: +41 (0)56 310 5297
>
> GPG Key ID = A46BEE1A
>
>
> Tommi Kajander, PhD
> Senior Scientist
> Structural Biology Program
> Institute of Biotechnology
> Biocenter 3, Viikinkaari
>  1 (PO
> Box 65)
> University of Helsinki
>
>
>
>
>
> --
>
> To unsubscribe from the CCP4BB list, click the following link:
> https://www.jiscmail.ac.uk/cgi-bin/webadmin?SUBED1=CCP4BB=1
>



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[ccp4bb] Normalization of B-factors

[Santhosh Gatreddy]

Hi all,

I have to compare the B-factors of three of my ligand bound structures of the 
same protein which were diffracted to 1.85, 1.98 and 2.01 A resolution.

Is it necessary to normalize the B-factors of these structures (dimer in ASU) 
before comparing them to understand the ligand induced stability.


Thanks for Your suggestions,

Santhosh


 








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Re: [ccp4bb] screw axes /system. absenses and phaser/MR solutions

[Kajander, Tommi A]

Hi Tim - There are reflections in all directions and only one clearly has 
systematic absenses. Although only few seem to be present in one direction (But 
thats the h00 direction here
+ all are present though) -phaser finds with same data P22121 as the strongest 
solution (few reflections are weak in that direction from below… but most not). 
 TZ-score diffrence is not
hure 13 vs 15.

P 2 2 21
  (   0,0,5): i/sigi =   20.8
  (   0,0,7): i/sigi =   19.6
  (   0,0,9): i/sigi =   21.0
  (   0,0,   11): i/sigi =1.9
  (   0,0,   13): i/sigi =   21.0
  (   0,0,   15): i/sigi =6.0
  (   0,0,   17): i/sigi =   20.8
  (   0,0,   19): i/sigi =0.9
  (   0,0,   21): i/sigi =9.0
  (   0,0,   23): i/sigi =1.0
  (   0,0,   25): i/sigi =4.9
  (   0,0,   27): i/sigi =   19.9
  (   0,0,   29): i/sigi =   20.9
  (   0,0,   31): i/sigi =   20.2
  (   0,0,   33): i/sigi =   19.1
  (   0,0,   35): i/sigi =   18.2
  (   0,0,   37): i/sigi =   13.1
  (   0,0,   39): i/sigi =   20.8
  (   0,0,   41): i/sigi =8.9
  (   0,0,   43): i/sigi =   12.7
  (   0,0,   45): i/sigi =2.5
  (   0,0,   47): i/sigi =0.8
P 21 2 2
  (   3,0,0): i/sigi =   31.2
  (   5,0,0): i/sigi =   25.2
  (   7,0,0): i/sigi =   21.0
  (   9,0,0): i/sigi =2.6
P 2 21 2
  (   0,5,0): i/sigi =0.7
  (   0,7,0): i/sigi =0.7
  (   0,9,0): i/sigi =0.7
  (   0,   11,0): i/sigi =1.1
  (   0,   13,0): i/sigi =1.0
  (   0,   15,0): i/sigi =1.5
  (   0,   17,0): i/sigi =0.7
  (   0,   19,0): i/sigi =0.7
  (   0,   21,0): i/sigi =0.7
  (   0,   23,0): i/sigi =0.8
  (   0,   25,0): i/sigi =0.7
  (   0,   29,0): i/sigi =0.7
  (   0,   31,0): i/sigi =0.8
  (   0,   33,0): i/sigi =0.7
  (   0,   35,0): i/sigi =0.7
  (   0,   37,0): i/sigi =0.9
  (   0,   39,0): i/sigi =0.7
  (   0,   41,0): i/sigi =0.7
  (   0,   43,0): i/sigi =0.7


Neither seem to refine now so have to work on it a bit more beyond this  - we 
see the same “conflict" with two different crystals - seems complicated - the 
other one processed much
better in P21  and seemed possibly twinned. Maybe the case here too.

Tommi





On Aug 9, 2018, at 10:29 AM, Tim Gruene 
mailto:tim.gru...@psi.ch>> wrote:

Dear Tommi,

did you check whether you collected any reflections at all that should
be absent for the second screw axis? If there are non - which could
easily happen with low resolution, incomplete data - pointless and XDS
might be conservative and not estimate the likelihood for the second
screw-axis.

MR, however, will pick up the space group with all data, and thus be
able to tell between P2212 and P22121 whether or not you recorded
reflections that are expected to be absent.

Best,
Tim

On 08/08/2018 04:29 PM, Kajander, Tommi A wrote:
Hi,
Any clues why the followting happens: pointless (and just looking at the XDS 
output) clearly tells there is one screw axis in P-ortorhombic (P2212)
yet phaser gives the best Z-scores in P22121. (...I suspect this may be to do 
with twinning - might be monoclinic twiined still though now processes very 
well in P222.)

If i run the mtz after XDSCONV (ie F2MTZ) via pointless (instead of directly 
after XDS) it also suggests this - but i suppose i am not suppose to run merged 
data via pointless.

Thanks for comments,

Tommi



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--
--
Paul Scherrer Institut
Tim Gruene
- persoenlich -
OSUA/204
CH-5232 Villigen PSI
phone: +41 (0)56 310 5297

GPG Key ID = A46BEE1A


Tommi Kajander, PhD
Senior Scientist
Structural Biology Program
Institute of Biotechnology
Biocenter 3, Viikinkaari 1 (PO Box 65)
University of Helsinki







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Re: [ccp4bb] screw axes /system. absenses and phaser/MR solutions

[Tim Gruene]

Dear Tommi,

did you check whether you collected any reflections at all that should
be absent for the second screw axis? If there are non - which could
easily happen with low resolution, incomplete data - pointless and XDS
might be conservative and not estimate the likelihood for the second
screw-axis.

MR, however, will pick up the space group with all data, and thus be
able to tell between P2212 and P22121 whether or not you recorded
reflections that are expected to be absent.

Best,
Tim

On 08/08/2018 04:29 PM, Kajander, Tommi A wrote:
> Hi,
> Any clues why the followting happens: pointless (and just looking at the XDS 
> output) clearly tells there is one screw axis in P-ortorhombic (P2212)
> yet phaser gives the best Z-scores in P22121. (...I suspect this may be to do 
> with twinning - might be monoclinic twiined still though now processes very 
> well in P222.)
> 
> If i run the mtz after XDSCONV (ie F2MTZ) via pointless (instead of directly 
> after XDS) it also suggests this - but i suppose i am not suppose to run 
> merged data via pointless.
> 
> Thanks for comments,
> 
> Tommi 
> 
> 
> 
> To unsubscribe from the CCP4BB list, click the following link:
> https://www.jiscmail.ac.uk/cgi-bin/webadmin?SUBED1=CCP4BB=1
> 

-- 
--
Paul Scherrer Institut
Tim Gruene
- persoenlich -
OSUA/204
CH-5232 Villigen PSI
phone: +41 (0)56 310 5297

GPG Key ID = A46BEE1A




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