Re: [ccp4bb] Semet in non-auxotrophic strains (summary)

- Protein crystallography

Main steps:

- Protein purification
- Crystallisation

Special:

- Programs for crystallography
- X-ray detectors

Basic tutorials:

   - Chemistry
   - Protein
   - Peptide
   - Amino Acids

Xtal community:

- CCP4BB

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

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From: Raja Dey deyraja1 {- at -} YAHOO {- dot -} CO {- dot -} IN
Date: 2008-11-03

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Subject: Re: Semet in non-auxotrophic strains (summary)
From: Jacob Keller j-keller2 {- at -} MD {- dot -} NORTHWESTERN {- dot -} EDU
Date: 2008-11-03

Hello all,

Prompted by Kay Diederichs, herein follows a summary of the responses
gleaned from the CCP4BB. The wiki has also been edited to include the
summary:

http://strucbio.biologie.uni-konstanz.de/ccp4wiki/index.php/Expression_of_SeMet_labeled_proteins

Summary:

1. Protocols for both auxotrophs and non-auxotrophs:
Doublié, S. (1997). Methods Enzymol. 276, 523-532

2. Two protocols for semet labelling:
http://www.doe-mbi.ucla.edu/local/protocols

3. Another non-auxotroph protocol. G. Fritz says this recipe could probably
be made in one vessel for immediate use, except for the aromatic aa's which
might dissolve better at basic pH:
http://strucbio.biologie.uni-konstanz.de/ccp4wiki/index.php/Expression_of_SeMet_labeled_proteins

4. Original literature source for met synthesis silencing (for use with
non-auxotrophs):
Van Duyne et al. J. Mol. Biol. 229, 105-124 (1993).

5. Autoinduction semet protocol (allegedly complicated recipe):
Studier FW (2005) Protein production by auto-induction in high density
shaking cultures. Protein Expr. Purif. 41, 207-234

Jacob Keller

*******************************************
Jacob Pearson Keller
Northwestern University
Medical Scientist Training Program
Dallos Laboratory
F. Searle 1-240
2240 Campus Drive
Evanston IL 60208
lab: 847.491.2438
cel: 773.608.9185
email: j-keller2@northwestern.edu
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CCP4bb navigation

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

Previous message:
Subject: nucplot
From: Raja Dey deyraja1 {- at -} YAHOO {- dot -} CO {- dot -} IN
Date: 2008-11-03

Next message:
Subject: m-3m vs. m-3
From: yanming Zhang shanmao62 {- at -} YAHOO {- dot -} COM
Date: 2008-11-03