HIS6 Tagged Int. Membrane Prot?

Richard Miller rmiller at bimcore.emory.edu
Tue Mar 14 00:54:19 EST 1995


   Has anyone tried to HIS6 tag an outer mitochondrial membrane protein
for purification by IDA chromatography?  I know that the mitochondrial
targeting sequence for my protein resides in the C-Terminus and that
manipulations of the N-Terminus do not affect targeting to the OMM.  I'm
using a yeast expression system which works great, but purification on a
large scale is proving to be a real problem. I'd like to attach a His6 tag
to the N-terminus to provide a means of easy purification, but I don't
know if the presence of detergents (VERY low concentration 0.05% Triton)
will affect binding of the protein to the IDA column.  If anyone has
attempted this (for any membrane protein) with either success or failure
please let me know.

Thanks in Advance,

Richard Miller
Department of Biochemistry
Emory University
Atlanta, GA



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