Re: [ccp4bb] exposing your Tev site

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CCP4bb <-- 1999 <-- November 1999 <-- 30 November 1999

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Subject: Re: recommendation for heavy atoms used in lithium sulfate and ammonium sulfate
From: Jeffrey {- dot -} Kieft {- at -} UCHSC {- dot -} EDU Jeffrey {- dot -} Kieft {- at -} UCHSC {- dot -} EDU
Date: 2007-04-20

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Subject: Re: recommendation for heavy atoms used in lithium sulfate and ammonium sulfate
From: artem {- at -} XTALS {- dot -} ORG artem {- at -} XTALS {- dot -} ORG
Date: 2007-04-20

Subject: Re: exposing your Tev site
From: Sebastien Violot svicri {- at -} IBMB {- dot -} CSIC {- dot -} ES
Date: 2007-04-20

Lukacs, Christine wrote:

> Quick question about Tev sites –
>
> We have a situation where our N-terminal TEV site is inaccessible to
> the protease. Looking at a close homolog, we realized that our first
> protein residue is already involved in a beta sheet. From peoples’
> experience – how FEW residues can we add between the cleavage site and
> the first protein residue in order to make the cleavage site
> accessible to the protease for efficient cleavage of the tag? (Tagged
> protein behaves well but crystallizes poorly, we are hoping that
> cleavage of the tag will improve things)
>
> Thanks for any anecdotes-
>
> ***Christine Lukacs*
>
> Roche
>
>
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Hi Christine,
According to our experience, a 3 amino-acids long linker (Ser-Gly-Ala)
between TEV recognition sequence and your protein of interest is enough
for efficient cleavage. Remember that after cleavage, this linker will
be part of your protein to be crystallized.
Good luck,

Seb

--
Dr Sebastien VIOLOT
Dep. Biologia Estructural
Institut de Biologia Molecular de Barcelona, CSIC
Parc Científic de Barcelona
Josep Samitier 1-5
08028 Barcelona
Spain

Phone +34 93 403 4957
Fax +34 93 403 4979
Email svicri@ibmb.csic.es
URL www.ibmb.csic.es

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CCP4bb <-- 1999 <-- November 1999 <-- 30 November 1999