Re: [ccp4bb] 2019 New Year’s resolutions of a cryo-EM newbie

[Marin van Heel]

Hello Jacob

I know it was a bit of a cynical joke and I know it does hurt ...  After
all my mother's Ellis Island records are dated 1923. Our last family
reunion was 15 years ago in Illinois and Missouri. My Mid-West raised uncle
was torpedoed off the coast of Cape Canaveral in 1942 transporting US
military equipment. And, then again, my youngest son is a US citizen. At
the same time, I also need to defend the integrity of scientific Canada in
name of my late Canadian sister who was an excellent biochemist. And no,
none of us were ever a member of the flat earth society (as far as I
know)!  ;)
Lets hope for a happier 2019!

Marin

On Wed, Jan 2, 2019 at 11:07 AM Keller, Jacob 
wrote:

> 6) I most sincerely hope that, if I stick to my five New Year’s
> resolutions and stop wasting the cryo-EM community’s time, my fellow
> Canadians will not extradite me to the fake-news country at our southern
> border.
>
>
>
>
>
> I know this is a joke, but we in the USA are not a fake news people, and
> please don’t punish us for the sins of our leadership--it’s painful for
> many of us.
>
>
>
> JPK
>
>
> --
>
> 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] Non-Cysteine modification with heavy atom

[Tom Peat]

Hello Chandra,

Soaking heavy atoms into crystals has a long and successful history (I use it 
pretty often and it works much of the time). If you need phase information and 
you have some crystals, I would certainly give it a try. There are many papers 
and books to give you methods to do this (good old Blundell and Johnson for 
one, Methods in Enzymology for another).
No reason to waste good native crystals if you still have some ;-)
Cheers, tom

Sent remotely

On 3 Jan 2019, at 6:43 am, Chandramohan Kattamuri 
<1c5b7cb6c764-dmarc-requ...@jiscmail.ac.uk>
 wrote:


Dear CCP4 members, Can someone suggest to me a methodology to covalently modify 
(non-cysteine) on component of a multicomponent complex with heavy atoms in 
order to achieve better phase determination?

Thanks in advance


Chandra
kattamuricha...@yahoo.com






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Re: [ccp4bb] Non-Cysteine modification with heavy atom

[Jesko Koehnke]

Dear Chandra,
A Million ways to die in the West (or when screening for HA derivatives).
But this has always worked for me:
https://www.cell.com/structure/fulltext/S0969-2126(00)00168-4
Just analyse ability to still form complex after modification.
Cheers,
Jesko

Am Mi., 2. Jan. 2019, 20:42 hat Chandramohan Kattamuri <
1c5b7cb6c764-dmarc-requ...@jiscmail.ac.uk> geschrieben:

>
> Dear CCP4 members, Can someone suggest to me a methodology to covalently
> modify (non-cysteine) on component of a multicomponent complex with heavy
> atoms in order to achieve better phase determination?
>
> Thanks in advance
>
>
> Chandra
> kattamuricha...@yahoo.com
>
>
>
>
> --
>
> 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] Non-Cysteine modification with heavy atom

[Artem Evdokimov]

Off the top of my head, assuming nothing much about your protein(s) - some
of this is only going to work if you can modify the protein(s) prior to
crystallization. This excludes the 'normal' heavy atom soaking type stuff
which can give rise to binding with covalent-like strength (while still
being noncovalent, technically):

Iodination of tyrosines (and histidines if pH is selected correctly)
Iodination or bromination of DNA/RNA if your complex contains nucleic acid
Introduction of heavy atom into a custom-made peptide (sky is the limit as
to what you can do with this) followed by peptide ligation (i.e. sortase
and the like) - requires modification and re-expression of your protein
construct of course
Reaction of Lys and N-term with suitably modified carboxylic acids (e.g.
para-iodo benzoic acid, para-mercuric derivatives and so on) via the use of
water soluble coupling agents or pre-activated acid derivatives
Reductive amination of Lys residues using aldehydes that bear heavy atom
labels (must be sure that the reducing agent does not knock the HA off,
which it jolly well can do)
Ditto with side chains of Asp and Glu, and the amine is modified with heavy
atom(s)
Introduction of unnatural amino acids (even bearing heavy atoms if desired)
by means of genetic manipulation (stop codon bypass, codon replacement etc.)
Nucleophilic attack on Serine (especially facile if you happen to have a
serine protease or something similar where serine is activated)

Also see: https://www.nature.com/articles/ncomms5740

Even though these methods are not necessarily selective with respect to
amino acids being modified, it is surprisingly often possible to only
modify a few (sometimes even one) specific positions in a protein due to
the selectivity imposed by protein surface environment

I am probably missing another 10-15 methods but gotta run :)

Artem

- Cosmic Cats approve of this message


On Wed, Jan 2, 2019 at 2:42 PM Chandramohan Kattamuri <
1c5b7cb6c764-dmarc-requ...@jiscmail.ac.uk> wrote:

>
> Dear CCP4 members, Can someone suggest to me a methodology to covalently
> modify (non-cysteine) on component of a multicomponent complex with heavy
> atoms in order to achieve better phase determination?
>
> Thanks in advance
>
>
> Chandra
> kattamuricha...@yahoo.com
>
>
>
>
> --
>
> 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] Non-Cysteine modification with heavy atom

[Chandramohan Kattamuri]

Dear CCP4 members, Can someone suggest to me a methodology to covalently modify 
(non-cysteine) on component of a multicomponent complex with heavy atoms in 
order to achieve better phase determination?

Thanks in advance


Chandra
kattamuricha...@yahoo.com






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Re: [ccp4bb] 2019 New Year’s resolutions of a cryo-EM newbie

[Artem Evdokimov]

My apologies for posting this on an open board :) I realize that this is
completely off topic - but it is a pretty painful topic that affects us all
regardless of academic or industrial affiliation, professional preferences
etc.

Artem

--- do not read below unless you want to :) ---

If an average citizen is asked whether they believe in fairies/unicorns or
in atoms (since you cannot see either* it's easy to get confused as to
which ones are real, I guess), they often pick fairies and unicorns. The
nature of belief as a great human fallacy does not cease to amaze me. Its
tendrils affect all of us, without exception.

Fake news is just the latest representation of the (utterly irrational)
power of belief. If one can convince millions that something is real (or
ethically/morally right) then fantasy may become 'real' for the duration of
the shared dream. To be fair, sometimes the dream is that of freedom,
liberty, equality, kindness, etc. but far too often it is the dream of the
dark and disturbing kind that leads to subjugation of self-determination,
oppression, whitewashing of atrocities, and violence.

Recent events in the political sphere simply illuminate some of the
abhorrent fantasies perpetrated by what amounts to nearly 50% of the
population. Mathematically speaking we are pretty close to a 'fake news
country' - especially since the 'dark dreamers' elicit more political power
than the rest of us. This did not begin two years ago. It will not end two
years from now.

Some thoughts on making things better (fairly basic for any practicing
scientist):

- always insist on proof, especially when common sense is at odds with
what's being 'sold'
- exercise passion and don't be afraid to get agitated

"Those who profess to favor freedom and yet depreciate agitation, are
people who want crops without ploughing the ground; they want rain without
thunder and lightning; they want the ocean without the roar of its many
waters." -- Frederick Douglass

* Yes, I realize that we can in fact see atoms with the right equipment.
This makes it even worse, all things considered...

- Cosmic Cats approve of this message


On Wed, Jan 2, 2019 at 8:07 AM Keller, Jacob 
wrote:

> 6) I most sincerely hope that, if I stick to my five New Year’s
> resolutions and stop wasting the cryo-EM community’s time, my fellow
> Canadians will not extradite me to the fake-news country at our southern
> border.
>
>
>
>
>
> I know this is a joke, but we in the USA are not a fake news people, and
> please don’t punish us for the sins of our leadership--it’s painful for
> many of us.
>
>
>
> JPK
>
>



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Re: [ccp4bb] 2019 New Year’s resolutions of a cryo-EM newbie

[Keller, Jacob]

6) I most sincerely hope that, if I stick to my five New Year’s resolutions 
and stop wasting the cryo-EM community’s time, my fellow Canadians will not 
extradite me to the fake-news country at our southern border.


I know this is a joke, but we in the USA are not a fake news people, and please 
don’t punish us for the sins of our leadership--it’s painful for many of us.

JPK



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[ccp4bb] Imaging support scientist and software scientist positions at Diamond light Source

[maria.harkiol...@diamond.ac.uk]

Dear CCP community,

I’d like to draw your attention to two vacancies we have currently at the X-ray 
tomography and super resolution fluorescence microscopy  beamline 
B24 at 
Diamond Light Source.

We are looking for a support 
scientist
 and a software 
scientist
 to join our team and help us develop and deliver cutting-edge microscopy 
applications for the life sciences.

Please forward this email to any interested parties and do let me know if you 
need further information.

The links are:
https://vacancies.diamond.ac.uk/vacancy/senior-support-scientist-b24-372441.html
https://vacancies.diamond.ac.uk/vacancy/software-data-analysis-scientist-373010.html

Wishing you all a happy and productive 2019!

All the very beast,

Maria

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