IUBio Biosequences .. Software .. Molbio soft .. Network News .. FTP

pMal-c2, modification of

Richard J. Dudley rdudley at pitt.edu
Fri Jan 28 08:54:40 EST 2000


Wolfgang--

After all these years that's the only vector you have?  My image is
shattered...

Do you have any vector with a T7 promoter you could use?  If so, you
could then use BL21 or a related strain to express your protein.

I have pMAL-c2X, and it looks like the NdeI site IS the start of MBC
translation, which lends a lot of credibility to the idea..  How big is
the protein you're trying to express?  Offhand, it looks perfectly
reasonable, but I've been caught in the perception vs. reality thing
before.  You'd essentially be turning pMAL-c2 into a pET vector, which
work on occasion for me.  On the upside, I have found that pMAL-c2X gives
a good protein yield in DH5a and TB1.

Good luck!

rich

Wolfgang Schechinger wrote:

> Hi all,
>
> I would like to express a His-tagged protein in E. coli. The only
> suitable vector I have at the moment, is pMal-c2.
> There is a unique restriction site (NdeI) right
> between the Ptac promotor and the start of malE translation.
> Thus I would cut out the whole maltose binding protein (and get rid
> of it since I don't need it anyway, because I have a His-tag) and
> insert my construct between this NdeI site and a the polylinker.
>
> Despite all this sounds quite reasonable to me, does anybody have
> any caveats, drawbacks or warnings to mention?
>
> All input is welcome!
>
> Wolfgang
>
> For those who don't know pMal-c2:
> pMal-c2 is a glucose inducible vector for procaryotic protein
> expression. When used normally, one has a fusion protein between
> maltose binding protein (malE) and the protein on interest; the
> bacterial lysate is purified by affinity purification using a amylose
> (maltose-like polymer) resin. Then the fusion protein is cleaved off
> proteolytically with factor Xa.
>
> Dr. Wolfgang Schechinger, Pathobiochemistry Dept.
> University of Tuebingen, Germany
> email: wolfgang.schechinger at med.uni-tuebingen.de
> wwWait: http://www.medizin.uni-tuebingen.de/~wgschech/start.htm
> -----
> *unsolicited mail is *NOT* appreciated, usual disclaimers apply
> -----
> Pressing ALT+F4 simultaneously enables you to read private messages
> ---

--
Richard J. Dudley (rdudley+ at pitt.edu)
Research Specialist V
Cystic Fibrosis Research Center
Dept. of Cell Biology and Physiology
University of Pittsburgh
http://www.cbp.pitt.edu
---> search BIONET archives at http://www.bio.net <---






More information about the Methods mailing list

Send comments to us at biosci-help [At] net.bio.net