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

2008-11-03 Thread Vangelis Christodoulou
UK' Subject: RE: [ccp4bb] Semet in non-auxotrophic strains (summary) By popular request I've placed our modified Se-Met method online: -Original Message- From: CCP4 bulletin board [mailto:[EMAIL PROTECTED] On Behalf Of Jacob Keller Sent: Monday, November 03, 2008 8:13 PM To: CCP4

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

2008-11-03 Thread Artem Evdokimov
Subject: RE: [ccp4bb] Semet in non-auxotrophic strains (summary) By popular request I've placed our modified Se-Met method online: -Original Message- From: CCP4 bulletin board [mailto:[EMAIL PROTECTED] On Behalf Of Jacob Keller Sent: Monday, November 03, 2008 8:13 PM To: CCP4BB@JISCMAIL.A

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

2008-11-03 Thread Artem Evdokimov
By popular request I've placed our modified Se-Met method online: -Original Message- From: CCP4 bulletin board [mailto:[EMAIL PROTECTED] On Behalf Of Jacob Keller Sent: Monday, November 03, 2008 8:13 PM To: CCP4BB@JISCMAIL.AC.UK Subject: Re: [ccp4bb] Semet in non-auxotrophic st

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

2008-11-03 Thread Jacob Keller
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 aux

Re: [ccp4bb] Semet in non-auxotrophic strains

2008-11-03 Thread Artem Evdokimov
4 bulletin board [mailto:[EMAIL PROTECTED] On Behalf Of Jacob Keller Sent: Monday, November 03, 2008 12:56 PM To: CCP4BB@JISCMAIL.AC.UK Subject: [ccp4bb] Semet in non-auxotrophic strains Dear Crystallographers, I have been using an E coli strain called Tuner (DE3) pLacI, which is not auxotrophic for

Re: [ccp4bb] Semet in non-auxotrophic strains

2008-11-03 Thread Jacob Keller
] *** - Original Message - From: Cynthia Kinsland To: Jacob Keller Cc: CCP4BB@JISCMAIL.AC.UK Sent: Monday, November 03, 2008 12:29 PM Subject: Re: [ccp4bb] Semet in non-auxotrophic strains You can express Se-Met protein in non-auxotrophs using metabolic inhibition

Re: [ccp4bb] Semet in non-auxotrophic strains

2008-11-03 Thread Cynthia Kinsland
You can express Se-Met protein in non-auxotrophs using metabolic inhibition (Doublie. Methods Enzymol. 276, 523-530, 1997 and Van Duyne et al. J. Mol. Biol. 229, 105-124, 1993). The Studier auto-induction paper (Studier, Prot. Exp. Purif. 41, 207-234, 2005) describes an auto-induction Se-Me

Re: [ccp4bb] Semet in non-auxotrophic strains

2008-11-03 Thread Arthur Laganowsky
Dear Jacob, You can find several protocols for Se-Met labeling at: http://www.doe-mbi.ucla.edu/local/protocols Good Luck, Art > Dear Crystallographers, > > I have been using an E coli strain called Tuner (DE3) pLacI, which is not > auxotrophic for methionine, nor is there an auxotrophic analog

Re: [ccp4bb] Semet in non-auxotrophic strains

2008-11-03 Thread Guenter Fritz
Hi Jacob, you can express SeMet labeled protein in almost any E.coli expression strain if you use a defined minimal medium. Here is a receipe that worked quite well in our hands: http://strucbio.biologie.uni-konstanz.de/ccp4wiki/index.php/Expression_of_SeMet_labeled_proteins HTH Guenter Dear Cr

[ccp4bb] Semet in non-auxotrophic strains

2008-11-03 Thread Jacob Keller
Dear Crystallographers, I have been using an E coli strain called Tuner (DE3) pLacI, which is not auxotrophic for methionine, nor is there an auxotrophic analogue available. It may be that my protein can be expressed in other strains, but this one works well right now, and it is usually better