4. protein monitoring
Dear Yogesha,
You can use the absorbance at 215 or 220 nm to follow your peptide
during purification. Compounds like DTT and EDTA can increase the
noise at those wavelengths, go to 220 nm if you have things like that
in your purification buffers.
Quantification o
Any protein will absorb at 230, and most UV detectors these days can
reach that, if you want to follow that with UV.
Together with SDS_PAGE of course, this should be good enough to go
ahead.
A.
On May 28, 2010, at 18:03, Sollepura Yogesha wrote:
Dear All,
I have expressed 30-40 aa region m
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 d
On Behalf Of
Sollepura Yogesha
Sent: Friday, May 28, 2010 12:04 PM
To: CCP4BB@JISCMAIL.AC.UK
Subject: [ccp4bb] protein monitoring
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
Hi Yogi,
You can see your peptide on a gel so why can't you "monitor" it by SDS-PAGE? A
little time consuming, yes, but then you have the extra benefit of also seeing if there
are contaminating proteins in your sample.
good luck,
Eric
__
Eric Larson, PhD
MSGPP Consor
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
