Chemical Cleavage of Proteins/Peptides

U12201 at uicvm.uic.edu U12201 at uicvm.uic.edu
Sat Mar 25 11:19:27 EST 1995


Try immobilizing enzymes on a support such as AminoLink or similar
from Pierce. This is what is used for Fab and Fab2 fragmentation
of antibodies. Immobilized enzymes give you controlled interaction
times and easy separations.
In article <3ksse1$hqo at nntp3.u.washington.edu>, kantola at u.washington.edu (Angeline Kantola) says:
>
>Greetings,
>
>I'm looking for references and/or advice about cleavage
>of proteins using chemical means. I have a couple of
>constructs of my protein of interest which use different
>(and efficient) purification schemes--His6 and a GST
>fusion, both N terminal. What I need is *just* my domain. Proteolysis
>using Factor Xa releases my protein from the GST fusion construct
>and then immediately digests my protein to teeny weeny bits.
>The His tag has persistant problems with dragging paramagnetic
>metals along with the protein into my NMR tube.
>
>CNBr is right out, since my sequence has several methionines.
>I've read about chemical cleavage after tryptophanes, but
>the presence of methionines seems to render that method much
>more complicated as well. Cleaving N terminal to cysteines
>using NTCB is more appealing; unfortunately I've yet to find
>a method to remove the cyclic end product at the N terminus.
>Anyone have a suggestion?
>
>I'm actively searching for a recent review of chemical means of
>protein cleavage but have had little success; suggestions
>for this are also appreciated.
>
>Much obliged,
>Angie Kantola
>kantola at u.washington.edu
>Graduate Student
>Department of Biochemistry
>University of Washington
>Seattle, WA  USA
>
>



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