Dear Marius,
I checked out the references, and it was mentioned that The zinc EXAFS
spectra of P. shermanii superoxide dismutase have shown that zinc can be
incorporated in the active center instead of the iron. (Eur. Biophys. J. *
24* , 243-250). And this example further aroused my concern
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
Since you sent your question to CCP4bb you presumably have crystals!
In that case an excellent way to check which metal atoms you have in
which sites is to collect datasets at a synchrotron at suitably
chosen wavelengths and look at the anomalous maps. For an example
see Acta Cryst. D62 (2006)
Dear Mr. Fritz,
Yes, the protein is not an E.coli protein! Instead, it was cloned from a
virus. And since it was a nonstructural viral protein, I thought it might be
appropiate to treat it as eukaryotic proteins.
E.coli system was quite different from eukaryotic ones, hence I was quite
cautious
Long time ago I had a superoxide dismutase that was
active with iron as well as manganese. No matter what
functional metal (Fe or Mn) was bound a
substantial fraction up to 1/3 of the molecules
had the (non-functional) Zinc in their metal
binding site (found by AAS and EXAFS).
Zinc is
As others have implied, quantifying metals in
metalloproteins is very challenging. In my experience, the principal
problems are (1) adventitious metal contamination, (2) accurate
measurement of protein concentration, and (3) weak metal binding.
Zinc and iron are ubiquitous microcontaminants,
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
