Protein-protein interaction of His-tag of pET14b?

Jared Head bijgh at zeus.bris.ac.uk
Mon Mar 3 09:18:23 EST 1997


Jimmy S.H. Tsang (jshtsang at hkucc.hku.hk) wrote:
: 
: Dear All,
:         I have recently fused a bacterial gene downstream of the His-tag 
: sequence of the pET14b from AMS Biotech.  The in vitro transcribed and 
: translated protein forms a complex which migrates much slower than the wild 
: type protein in a native PAGE.  Did anyone experience a similar 
: 'multimerisation' problem?  The same gene cloned into pET19b produced a 
: protein similar to that of native protein, i.e. no 'multimerisation'.  Any 
: suggestion and comment will be appreciated.

We're just finding the same thing.  It seems possible that more than one
protein are chelating around one nickel atom.  When we cleaved the His.Tag
off we made our protein monomeric, and we also found dtt to have the same
effect, presumably by reducing the nickel ions.

Jared

-- 
Jared Head     at the Department of Biochemistry, University of Bristol

"A computer lets you make more mistakes faster than any invention in human
     history - with the possible exceptions of handguns and tequila." 
                                                   Mitch Ratliffe



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