Re: [ccp4bb] Different conformation of chains

[Petr Kolenko]

Dear Shine.star1609,
Your structural observation may be really different from the original one. 
There are several things to do now:


  1.  Check whether the same interpretation could be possible in the original 
structure. Are there maps or the original diffraction data? You could also 
reprocess them.
  2.  Read the published methods carefully and compare your protocol to the 
published one. The different structure could be interpreted based on the 
differences.
  3.  Have you used the same ligand? If not, both the observations are valid 
and possible. Analyze the interactions. This would definitely be an additional 
information for your paper.
  4.  Different conformations in “the same but opposite” chains are possible 
and observed also in other structures. Are there more structures of this 
protein?
  5.  Is there a possibility to contact the original authors?

I am sure that there will be more comments soon. 
Best regards,
Petr


Od: CCP4 bulletin board  za uživatele S
Odesláno: Thursday, October 21, 2021 2:52 AM
Komu: CCP4BB@JISCMAIL.AC.UK
Předmět: [ccp4bb] Different conformation of chains

Dear All,

I am working with a protein, the structure of which is already published in the 
PDB (2 chains in asu) The published structure has a break of around 20 residues.

I have crystallized the same protein and saw some density near the gap (20 
residues break).
I could build 3-4 residues in ChainA and 8 residues in ChainB (in the 20 
residue break region). After refinement these showed good density and b factor.
But the issue is that in ChainA, it goes straight. While in ChainB, the 8 
residues converge into a loop and goes to the other side.

The protein is a monomer in solution, is it possible to have different 
conformation in different chains that is completely opposite.
Also the 8 residues (ChainB) that is build now occupies the site where ligand 
was placed in the published structure. I am not sure how to interpret this 
(since in ChainA that site is unoccupied as contrast to ChainB).

Any thoughts/suggestions will be really helpful.

Thanks you




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Re: [ccp4bb] Different conformation of chains

[Pavel Afonine]

Hi,

if I understood your question correctly, then one of 13 scenarios described
here:

http://phenix-online.org/phenixwebsite_static/mainsite/files/newsletter/CCN_2015_07.pdf#page=12

fits your situation and you should be able to handle it in refinement all
right.

For more specific advice please feel free to contact me directly (off list)
with relevant files.

Good luck!
Pavel

On Wed, Oct 20, 2021 at 5:53 PM S  wrote:

> Dear All,
>
> I am working with a protein, the structure of which is already published
> in the PDB (2 chains in asu) The published structure has a break of around
> 20 residues.
>
> I have crystallized the same protein and saw some density near the gap (20
> residues break).
> I could build 3-4 residues in ChainA and 8 residues in ChainB (in the 20
> residue break region). After refinement these showed good density and b
> factor.
> But the issue is that in ChainA, it goes straight. While in ChainB, the 8
> residues converge into a loop and goes to the other side.
>
> The protein is a monomer in solution, is it possible to have different
> conformation in different chains that is completely opposite.
> Also the 8 residues (ChainB) that is build now occupies the site where
> ligand was placed in the published structure. I am not sure how to
> interpret this (since in ChainA that site is unoccupied as contrast to
> ChainB).
>
> Any thoughts/suggestions will be really helpful.
>
> Thanks you
>
>
> --
>
> To unsubscribe from the CCP4BB list, click the following link:
> https://www.jiscmail.ac.uk/cgi-bin/WA-JISC.exe?SUBED1=CCP4BB=1
>



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[ccp4bb] Different conformation of chains

[S]

Dear All,

I am working with a protein, the structure of which is already published in
the PDB (2 chains in asu) The published structure has a break of around 20
residues.

I have crystallized the same protein and saw some density near the gap (20
residues break).
I could build 3-4 residues in ChainA and 8 residues in ChainB (in the 20
residue break region). After refinement these showed good density and b
factor.
But the issue is that in ChainA, it goes straight. While in ChainB, the 8
residues converge into a loop and goes to the other side.

The protein is a monomer in solution, is it possible to have different
conformation in different chains that is completely opposite.
Also the 8 residues (ChainB) that is build now occupies the site where
ligand was placed in the published structure. I am not sure how to
interpret this (since in ChainA that site is unoccupied as contrast to
ChainB).

Any thoughts/suggestions will be really helpful.

Thanks you



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[ccp4bb] tech and postdoc position at Brown University

[Deaconescu, Alexandra]

Hello:

Two positions are available in my lab at Brown University. Please see below
and feel free to email me for more information.

*POSTDOCTORAL AND TECHNICIAN POSITIONS*

*at Brown University, USA*



*The Deaconescu Lab* at Brown University has one opening for a postdoctoral
researcher and one for a technician. The laboratory's interests lie
primarily in stress responses, with particular emphasis on responses to DNA
damage. We utilize a combination of biochemical, biophysical and structural
techniques. (e.g. X-ray crystallography, electron microscopy). Examples of
recent work are: Schwartz et al, *Protein Science *(2021), Brugger et
al, *Nature
Communications *(2020), Dorich et al, *Genes & Development *(2019), Vemu et
al. *Science *(2018), Le TT et al. *Cell* (2018), Szyk et al, *Cell* (2014).

*A successful postdoctoral candidate* should have an established
track-record of publications in peer-reviewed journals; solid experience
with protein biochemistry (their purification and assay development). We
have a variety of projects available at different stages of development,
including some that involve drug design. Prior knowledge of crystallography
and/or single-particle electron microscopy is highly desirable. Must be
highly motivated and work well independently as well as in a team.
Excellent spoken and written English are required. Newly minted PhDs are
encouraged to apply.


*A successful technician candidate* should preferably have  a degree in
chemistry/biological sciences and at least one year experience at the
bench, at the minimum experience with molecular biology, and ideally, also
with protein purification and crystallization.


*Interested candidates should send a CV, a one page research experience
summary and contact information for three references to *
alexandra_deaconescu[at]brown.edu. Please use “postdoctoral candidate”
or “technician candidate" as a subject.


Salary and starting date are negotiable. Brown University, an Ivy League
school ranked as 5th best in the US by the Wall Street Journal, is located
in Providence, Rhode Island less than one hour away by train from Boston.
Rhode Island provides plenty of opportunities for outdoor recreation with
beautiful beaches and sailing and we offer a friendly and supportive lab
environment.



Lab webpage: https://deaconesculab.com



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[ccp4bb] Research Staff Position at Baylor College of Medicine

[Tsai, Francis T.F.]

STAFF MEMBER / ASSOCIATE

A full-time staff position is available in the Advanced Technology Core for 
Macromolecular X-ray Crystallography at Baylor College of Medicine. The mission 
of the X-ray core is to assist academic researchers and physician-scientists in 
finding a structure solution of their macromolecule of interest to develop 
novel treatments against unmet human diseases. The successful applicant will 
assist with the robotic screening, imaging, evaluation and optimization of 
macromolecular crystals of protein and protein-drug complexes. Experience with 
protein expression and purification is an advantage. Salary is competitive and 
in addition to a generous fringe benefit package. The position is ideally 
suited for a graduate with an M.S. or B.S. in a natural science with prior wet 
lab experience (this is not a solicitation for graduate student trainees). The 
core laboratory is located on the main BCM campus that provides a safe and 
secure environment for basic research. Social distancing and other safety 
precautions have been implemented and are maintained at all times.

Primary duties and responsibilities:

  *   Assist with crystallization screening setups using a liquid handling 
robot.
  *   Assist with imaging and recording of crystallization screening results.
  *   Prepare biological buffers and chemical stock solutions.
  *   Perform experiments in protein biochemistry.
  *   Prepare figures and assist in writing reports related to experiments 
performed.
  *   Maintain general lab environment (i.e., organization and cleanliness of 
the lab).
  *   Maintain and ensure compliance with biosafety regulations.

Preferred qualifications:

  *   Required: A Master's degree or Bachelor's with 2 years of related lab 
experience.
  *   Desired: Prior experience with protein purification or biochemistry 
techniques.
  *   Ability to keep accurate records of work performed is essential.
  *   Strong organizational and communication skills.
  *   Strong work ethic, including ability to prioritize tasks, use good 
judgment and complete duties.
  *   Able to work independently and as a team.
  *   Willingness to learn new techniques.
To apply, please send a cover letter and current CV with contact information 
for two referees to the Director of the X-ray core laboratory, Dr. Sukyeong Lee 
(atc-xrayc...@bcm.edu). Highly motivated 
researchers with excellent communication skills are encouraged to apply. Baylor 
College of Medicine is an equal opportunity employer.




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[ccp4bb] Protein Purification jobs at UCB (Slough, UK)

[Leysen Seppe]

Dear all

Do you enjoy purifying challenging proteins?  Do you want to support antibody 
and small molecule drug discovery projects to transform patients' lives for the 
better?
Then consider applying for these roles in our Protein Production group:

Senior Scientist - Protein Biochemist job in Slough, Berkshire, United Kingdom 
| Research & Development jobs at 
UCB

Research Scientist - Protein Biochemist job in Slough, Berkshire, United 
Kingdom | Research & Development jobs at 
UCB

Instructions to apply can be found on the website.

Best regards

Seppe Leysen
Principal Scientist
Protein Structure & Biophysics
UCB


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other than the intended recipients is prohibited and may be illegal. If you are 
not an intended recipient, please immediately inform the sender and return the 
electronic mail and its attachments and destroy any copies which may be in your 
possession. UCB screens electronic mails for viruses but does not warrant that 
this electronic mail is free of any viruses. UCB accepts no liability for any 
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[ccp4bb] Reminder: CBMS Lecture Series - October 20 - Lee Makowski

[Stojanoff, Vivian]

Please join us ………..



Molecular Events in the Progression of Alzheimer’s Disease



Lee Makowski



Northeastern University



WEDNESDAY, October 20, 13:30 (EST)



Register in advance for this meeting:

https://bnl.zoomgov.com/meeting/register/vJIscuqgrD0vG8SVmZ49QwTrS8eGSZrs-1U



Abstract:

Fibrillar aggregates of Abeta peptides and tau protein are defining features of 
Alzheimer's disease (AD) but the role these structures play in the etiology of 
disease remains uncertain. Outstanding questions remain as to the distribution 
of polymorphs between and within cases.  We are using x-ray scanning 
microdiffraction on histological sections of human brain tissue in order to map 
the distribution and arrangement of fibrillar aggregates of these proteins in 
plaques and tangles.  The central hypothesis of the work is that the spatial 
distribution of structural polymorphs in brain tissue will provide important 
clues as to how fibrils contribute to disease.  Our goals are to assess whether 
or not different fibrillar polymorphs spread by a prion-like process during 
disease progression and to produce data that will provide insight into the 
structural basis by which different fibrillar strains are associated with 
different disease subtypes.






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[ccp4bb] LMB Kendrew Lecture on AlphaFold

[Jan Löwe]

Dear all,

I thought you might be interested to know that the recent John Kendrew 
Lectures 
 
at the MRC Laboratory of Molecular Biology, given by Demis Hassabis and 
John Jumper from DeepMind on AI in science and on the implementation and 
performance of AlphaFold2 have now been posted on YouTube:


https://www.youtube.com/watch?v=sm-VkgVX-2o 



https://www.youtube.com/watch?v=jTO6odQNp90 



With best wishes,

Jan




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[ccp4bb] Postdoctoral position in FBDD in Cambridge UK

[Marko Hyvonen]

Dear colleagues,

We have an opening in our group at the Department of Biochemistry, University 
of Cambridge,  for a postdoctoral researcher for 12 months to work on an 
exciting drug discovery project with Alborada Drug Discovery Institute 
(https://cambridge-ddi.alzheimersresearchuk.org/) to support their inhibitor 
design process with fragment-based approaches and with structural analyses.

We'd hope to find someone with expertise in this area already, be it in 
crystallography (we hope to run a large fragment screen against the target), 
NMR (ligand-observe fragment screens), biophysics (ITC, SPR, ...)  and/or 
chemistry, but significant support around as well.  Plenty of work to be done 
to understand how the inhibitors work, so experience in mechanistic analysis of 
proteins would be very useful.

Details from University of Cambridge application portal: 
https://www.jobs.cam.ac.uk/job/31927/

Happy to receive informal queries by email as well.

Please do spread the word towards any potential candidates.

Best, Marko


Marko Hyvonen
Department of Biochemistry
University of Cambridge
https://hyvonen.bioc.cam.ac.uk
@HyvonenGroup
mh...@cam.ac.uk




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[ccp4bb] postdoc position available in the lab of Prof. Eddy Arnold (Rutgers University, NJ, USA)

[F. Xavier Ruiz]

Dear All,

We have an immediate opening for a skilled post-doctoral researcher in the 
laboratory of Prof. Eddy Arnold at Rutgers University (NJ, USA). The aim of the 
project is to study the structure and function of human viral proteins, mainly 
polymerases from HIV-1, SARS-CoV-2 and other RNA viruses posing a pandemic 
threat, through X-ray crystallography and cryo-electron microscopy.

For more information about the position, please see the full advert: 
https://jobs.rutgers.edu/postings/145198 
Interested candidates should apply via the above link. For informal inquiries, 
please feel free to contact me directly.

Best wishes,

F. Xavier RUIZ
Assistant Research Professor
Arnold Laboratory Center for Advanced Biotechnology and Medicine Rutgers 
University
https://cabm.rutgers.edu/research/arnold-lab



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Re: [ccp4bb] am I doing this right?

[Ian Tickle]

Hi Kay

Can I just confirm that your result Iobs=0.01 sigIobs=0.01 is the estimate
of the true average intensity *per pixel* for a patch of 100 pixels?  So
then the total count for all 100 pixels is 1 with variance also 1, or in
general for k observed counts in the patch, expectation = variance = k+1
for the total count, irrespective of the number of pixels?  If so then that
agrees with my own conclusion.  It makes sense because Iobs=0.01
sigIobs=0.01 cannot come from a Poisson process (which obviously requires
expectation = variance = an integer), whereas the total count does come
from a Poisson process.

The difference from my approach is that you seem to have come at it via the
individual pixel counts whereas I came straight from the Agostini result
applied to the whole patch.  The number of pixels seems to me to be
irrelevant for the whole patch since the design of the detector, assuming
it's an ideal detector with DQE = 1 surely cannot change the photon flux
coming from the source: all ideal detectors whatever their pixel layout
must give the same result.  The number of pixels is then only relevant if
one needs to know the average intensity per pixel, i.e. the total and s.d.
divided by the number of pixels.  Note the pixels here need not even
correspond to the hardware pixels, they can be any arbitrary subdivision of
the detector surface.

Best wishes

-- Ian


On Tue, 19 Oct 2021 at 12:39, Kay Diederichs 
wrote:

> James,
>
> I am saying that my answer to "what is the expectation and variance if I
> observe a 10x10 patch of pixels with zero
> counts?" is Iobs=0.01 sigIobs=0.01 (and Iobs=sigIobs=1 if there is only
> one pixel) IF the uniform prior applies. I agree with Gergely and others
> that this prior (with its high expectation value and variance) appears
> unrealistic.
>
> In your posting of Sat, 16 Oct 2021 12:00:30 -0700 you make a calculation
> of Ppix that appears like a more suitable expectation value of a prior to
> me. A suitable prior might then be 1/Ppix * e^(-l/Ppix) (Agostini §7.7.1).
> The Bayesian argument is IIUC that the prior plays a minor role if you do
> repeated measurements of the same value, because you use the posterior of
> the first measurement as the prior for the second, and so on. What this
> means is that your Ppix must play the role of a scale factor if you
> consider the 100-pixel experiment.
> However, for the 1-pixel experiment, having a more suitable prior should
> be more important.
>
> best,
> Kay
>
>
>
>
> On Mon, 18 Oct 2021 12:40:45 -0700, James Holton  wrote:
>
> >Thank you very much for this Kay!
> >
> >So, to summarize, you are saying the answer to my question "what is the
> >expectation and variance if I observe a 10x10 patch of pixels with zero
> >counts?" is:
> >Iobs = 0.01
> >sigIobs = 0.01 (defining sigIobs = sqrt(variance(Iobs)))
> >
> >And for the one-pixel case:
> >Iobs = 1
> >sigIobs = 1
> >
> >but in both cases the distribution is NOT Gaussian, but rather
> >exponential. And that means adding variances may not be the way to
> >propagate error.
> >
> >Is that right?
> >
> >-James Holton
> >MAD Scientist
> >
> >
> >
> >On 10/18/2021 7:00 AM, Kay Diederichs wrote:
> >> Hi James,
> >>
> >> I'm a bit behind ...
> >>
> >> My answer about the basic question ("a patch of 100 pixels each with
> zero counts - what is the variance?") you ask is the following:
> >>
> >> 1) we all know the Poisson PDF (Probability Distribution Function)
> P(k|l) = l^k*e^(-l)/k!  (where k stands for for an integer >=0 and l is
> lambda) which tells us the probability of observing k counts if we know l.
> The PDF is normalized: SUM_over_k (P(k|l)) is 1 when k=0...infinity is 1.
> >> 2) you don't know before the experiment what l is, and you assume it is
> some number x with 0<=x<=xmax (the xmax limit can be calculated by looking
> at the physics of the experiment; it is finite and less than the overload
> value of the pixel, otherwise you should do a different experiment). Since
> you don't know that number, all the x values are equally likely - you use a
> uniform prior.
> >> 3) what is the PDF P(l|k) of l if we observe k counts?  That can be
> found with Bayes theorem, and it turns out that (due to the uniform prior)
> the right hand side of the formula looks the same as in 1) : P(l|k) =
> l^k*e^(-l)/k! (again, the ! stands for the factorial, it is not a semantic
> exclamation mark). This is eqs. 7.42 and 7.43 in Agostini "Bayesian
> Reasoning in Data Analysis".
> >> 3a) side note: if we calculate the expectation value for l, by
> multiplying with l and integrating over l from 0 to infinity, we obtain
> E(P(l|k))=k+1, and similarly for the variance (Agostini eqs 7.45 and 7.46)
> >> 4) for k=0 (zero counts observed in a single pixel), this reduces to
> P(l|0)=e^(-l) for a single observation (pixel). (this is basic math; see
> also §7.4.1 of Agostini.
> >> 5) since we have 100 independent pixels, we must multiply the
> individual PDFs to get the overall PDF f, and also