Hi Jack,
just a famous example:
The LIM region of a presumptive Caenorhabditis elegans transcription
factor is an iron-sulfur- and zinc-containing metallodomain.
Li PM, Reichert J, Freyd G, Horvitz HR, Walsh CT.
Proc Natl Acad Sci U S A. 1991 Oct 15;88(20):9210-3.
and
Cysteine-rich LIM domains of LIM-homeodomain and LIM-only proteins
contain zinc but not iron.
Archer VE, Breton J, Sanchez-Garcia I, Osada H, Forster A, Thomson AJ,
Rabbitts TH.
Proc Natl Acad Sci U S A. 1994 Jan 4;91(1):316-20.
another is
Free Radic Biol Med. 1999 Jul;27(1-2):193-202.
Expression, purification, and biochemical characterization of SAG, a
ring finger redox-sensitive protein.
Swaroop M, Bian J, Aviram M, Duan H, Bisgaier CL, Loo JA, Sun Y.
My own unpublished example is a DAG binding domain containing partially
a nice 3Fe-4S cluster instead of a Zn.
A brownish instead of a colourless protein solution is a good hint.
Best,
Guenter
Dear Guenter,
Would you mind sending me or posting on the ccp4bb the references to the cases
you referred to in your message about eukaryotic Zn-proteins expressed in
E.coli containing Fe instead of Zn?
Thanks.
Jack Tanner
-----Original Message-----
From: CCP4 bulletin board on behalf of Guenter Fritz
Sent: Mon 8/6/2007 1:24 AM
To: CCP4BB@JISCMAIL.AC.UK
Subject: [Junk Mail] Re: [ccp4bb] Zinc or Iron binding protein, that is a
question!
Hi Xuan,
I guess your protein is not an E.coli protein. There are several
examples that eukaryotic Zn-proteins expressed in E.coli contain Fe
instead of Zn. I am sceptic whether IMAC with different metal ions will
give the solution of the problem. If you really want to get information
on the metal ion binding properties you will have to do some matallo
biochemistry: preparing apo protein, reconstitution with metal ions,
UV-Vis spectroscopy, EPR would be great, ...
Dear Sir or Madam,
The ICP-ES results indicated that 1 molar my protein purified from
E.coli Origami(DE3) contained about a half molar Zinc and nearly a
quarter molar Iron (whether II or III was not available). The protein
carried a MBP tag on the N-terminal and the situation was similar with
or without His tag at the C terminal. I want to determine whether my
protein really bind Zinc or Iron. Does anyone have any experience
about such problems?
Specifically, now I want to compare the binding efficiency on various
IMAC, i.e. 50mM ZnSO4, FeSO4, Fe2(SO4)3, NiSO4(control), or
CuSO4(control). However, considering the instability of Fe(II) in
solution, the design still seemed problematic.
Sincerely,
Xuan Yang
National Laboratory of Biomacromolecules and
Center for Infection and Immunity,
Institute of Biophysics,
Chinese Academy of Sciences,
Room 1617, 15 DaTun Road,Chaoyang District,
Beijing, China, 100101
Tel: 86-10-64884329
Academic email: ya...@moon.ibp.ac.cn <mailto:ya...@moon.ibp.ac.cn>
We will either find a way or make one.
--
***********************************
Priv.Doz.Dr. Guenter Fritz
Fachbereich Biologie
Sektion Naturwissenschaften
Universitaet Konstanz
http://www.biologie.uni-konstanz.de/fritz
Universitaetsstrasse 10
Postfach M665
D-78457 Konstanz
e-mail: guenter.fr...@uni-konstanz.de
Phone Office: +49-(0)7531 88 3205
Phone Lab : +49-(0)7531 88 3733
Fax: +49-(0)7531 88 2966
