n terminal deblocking of protein in gel or pvdf membrane

Houthaeve Tony houthaeve at embl-heidelberg.de
Fri May 19 02:50:43 EST 1995


<barbara.valsasina at itner.pharmacia.se> wrote:
>      sometimes I have to deal with proteins electroblotted onto pvdf 
>      membrane or in sds page that are impossible to sequence because they 
>      are blocked at the N terminal end .
>      I usually try CNBr cleavage but after that I have problems in the 
>      extraction of obtained peptides.
>      I would like to know if anybody has any experience in deblocking 
>      blotted proteins as for example with methanolysis or in extraction 
>      from pvdf (I use Problott membranes - from Applied Biosystems).
 

Just forget it. You have, when having not enough material, hardly any
chance to deblock !
What do you exactly want ?
Straightforward, I'd say, perform a peptide map with trypsin and check 
on MALDI. Then fish out the peptide + eventually the blocking group.

Tony H.

EMBL
Heidelberg




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