Hello Claudia,
have you thought of, instead of rules, using restricted proteolysis combined
with N-terminal sequencing in order to better estimate a good position at least
for the N-terminal part of your protein?
Tim
On Thu, Mar 11, 2010 at 09:26:53AM +0100, Anastassis Perrakis wrote:
> Hi -
>
> I do not think there are "rules". There is a logic, to never truncate
> within secondary structure elements, and to prefer to delete regions
> that have a high probability to be disordered.
>
> With excuses for the non-ccp4 plugin, you can use our web server to help
> you design these.
>
> http://xtal.nki.nl/ccd/Welcome.html
>
>
> A.
>
>
> On Mar 10, 2010, at 12:45, Claudia Scotti wrote:
>
>>
>> Dear All,
>>
>> I'd like to espress a protein for crystallisation as a whole, but also
>> to try separate expression of its two domains.
>>
>> Although prediciting domain boundaires is not difficult, I know that
>> it is worthwhile testing several constructs to improve espression in
>> soluble form. In several works I've seen that the choice is to keep or
>> give up some aminoacids (for example 4 or 5) at the N and/or C-termini
>> and to test them to see which expresses best.
>>
>> I was wondering if there are defined "rules" to choose these variants
>> and/or to modify the N and C termini of a protein of interest when it
>> is not important to keep them "true" (=native) in order to optimise
>> expression levels.
>>
>> Is there any good reference about this, please?
>>
>> Personal experiences are welcome.
>>
>> Many thanks,
>>
>> Claudia
>>
>>
>> Claudia Scotti Dipartimento di Medicina Sperimentale Sezione di
>> Patologia Generale Universita' di Pavia Piazza Botta, 10 27100 Pavia
>> Italia Tel. 0039 0382 986335/8/1 Facs 0039 0382 303673
>>
>>
>>
>> Hotmail: Free, trusted and rich email service. Get it now.
>
> P please don't print this e-mail unless you really need to
> Anastassis (Tassos) Perrakis, Principal Investigator / Staff Member
> Department of Biochemistry (B8)
> Netherlands Cancer Institute,
> Dept. B8, 1066 CX Amsterdam, The Netherlands
> Tel: +31 20 512 1951 Fax: +31 20 512 1954 Mobile / SMS: +31 6 28 597791
>
>
>
>
--
--
Tim Gruene
Institut fuer anorganische Chemie
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D-37077 Goettingen
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