beta galactosidase purification

Bipin K Dalmia bipin at
Thu May 12 17:21:15 EST 1994

In article <1994May9.102353.1 at>,
 <rfeldber at> wrote:
>	I have been asked to put together a section of a teaching lab which
>will demonstrate enzyme purification/characterization using the beta
>galactosidase of E. coli. Does anyone already have a simple, fool-proof
>protocol for partial purification of this enzyme? I was thinking of asking
>different groups of students to use different protocols and then compare
>specific activities and gel patterns. Any suggestions would be greatfully

the method of steers (i can dig up the reference if you don't have it)
has always worked for me. just buy beta-d-thiogalactopyranoside agarose
from sigma, pack it in a column, run the mixture of proteins with b-gal
at around pH 7. b-gal will bind very specifically. wash off everything
else and elute with 100 mM sodium borate pH 10.00. minimize the exposure
to pH 10 by immediately dialyzing or do what i used to do, elute in a
equal volume of 1 M tris-hcl pH 7.0. this would dilute your enzyme but
would recover almost 100 % of the activity. you may already know this,
but b-gal needs 10 mM MgCl2 and 10 mM 2-mercaptoethanol in all buffers
to be stable.


bipin k. dalmia               the other night i was lying on my bed, looking
bipin at             up at the beautiful stars, and i said to myself, 
n2.bkd at     'where the F*CK is my ROOF !!'

More information about the Methods mailing list