You have to be careful with the Bradford detection assays because some
of them have a lower detection limit of 3-5 kDa and may not work for
your large peptide.
For FPLC you may be able to detect your protein using absorption at 215
nm which detects the peptide bond. Just choose a buffer that does not
absorb in this range.
Ursula
On 5/28/10 9:03 AM, Sollepura Yogesha wrote:
Dear All,
I have expressed 30-40 aa region my protein fused to GST.
I subjected it to precision protease cleavage. On the gel I can see
the band.
When I looked for ProtParam in expasy it shows that my peptide doesn't have *Extinction coefficients as* " there are no Trp, Tyr or Cys in the region considered, your protein should not be visible by UV spectrophotometry."
I need to separate GST from the cleavage mixture.
How can I monitor my peptide during FPLC and after that.
AA composition is Ala (A) 8, Arg (R) 2, Asn (N) 3, Asp (D)
2, Gln (Q) 2, Glu (E) 2, Gly (G) 4, Ile (I) 1, Leu (L)
4, Lys (K) 4, Phe (F) 2, Pro (P) 2, Ser (S) 5, Thr (T)
5, Val (V) 3.
I am looking for some suggestions
Thanks in advance
Yogi
--
Ursula Schulze-Gahmen, PhD.
QB3, Tjian Lab
MCB, 16 Barker Hall #3204
University of California Berkeley
Berkeley, CA 94720-3204
Phone: (510) 642 8258
[email protected]
