Dear Sylvie,
Any maleimide-activated KLH is going to react with the sulfhydryl
group of any cysteine, whether it is at the C- or N-terminus. You
probably would get coupling at both your C-terminal cysteines. Since you
appear to have only one free amino group, at the N-terminus, you probably
want to use something like glutaraldehyde to couple your peptide to your
carrier protein, after attaching something as a spacer, some glycines, for
example, or some epsilon amino hexanoic acid.
Hope this helps.
Angela C. Murphy
On Tue, 31 Oct 2000, SSPEQ wrote:
> Hi everyone,
>
> When I couple my peptides to KLH, I like to use the NEM-activated KLH and
> add a cysteine either at the N or C terminus of the peptide depending on
> the location of the peptide in the protein.
>
> For a particular peptide (see below), which is at the C-terminus end of the
> protein, I think it would be better to couple the KLH at the N-terminus of
> the peptide. But, this particular peptide has already two cysteines at the
> C-terminus. Would you use those Cys or you would add a Cys at the
> N-terminus or else?
>
> I wanted to find a reagent specific for N-terminus but that would not react
> with the cysteine, but by looking in the Pierce catalog, it seems they will
> all react with the cysteine also .
>
> Here is the sequence RXXXXXXXXXCCG where X are D, G, L, P,S,T, V.
>
> I would appreciate any tips on the strategy that you would use!
>
> Sylvie
>
>
> Sylvie Bourassa Ph.D.
> Service Proteomique de l'Est du QuÈbec (SPEQ)
> Anciennement: Service de Sequence de Peptides de l'Est du Quebec (SSPEQ)
> CHUL Research Center Room 9700
> 2705, Laurier blvd
> Ste-Foy, Quebec
> Canada G1V 4G2
> phone: 418-656-4141 ext 7088 /418-654-2267
> fax: 418-654-2159
> Email: sspeq@crchul.ulaval.ca
>
> http://www.crchul.ulaval.ca/crchul/fr/serv/sspeq.asp
>
>
*******************************************
Angela C. Murphy, Chemist
Lab. of Cell Biology, NHLBI, NIH
3 Center Drive, MSC 0301, Rm. B1-22
9000 Rockville Pike
Bethesda, MD 20892-0301 USA
tel.: (301) 496-2324
fax: (301) 402-1519
email: acmurphy@helix.nih.gov
*******************************************
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