Measuring peptide concentration

Artem Evdokimov AEVDOKIMOZ at cinci.rr.com
Tue Mar 19 19:28:45 EST 2002


Yes, Cys has an absorption peak there. Depending on the state - CyS-SyC is
different from two Cys !

No worries :)

A.G.E.

"Emir Khatipov" <khatipovNO at NOuchicago.edu> wrote in message
news:xjOl8.347$s4.26393 at news.uchicago.edu...
> So these are cysteines that give some fluorescence to my non-aromatic
> peptides! It's never too late to learn. Thanks for opening my eyes a
little
> bit wider, Artem :-)
> Cheers,
> Emir
>
> "Artem Evdokimov" <AEVDOKIMOZ at cinci.rr.com> wrote in message
> news:GBxl8.166129$Hu6.38723197 at typhoon.neo.rr.com...
> > I disagree on one point : Expasy server does NOT calculate extinciton if
> you
> > don't have certain types of AA's in your sequence.
> > For example 'AAAAAA' results in
> >
> > -----------
> > Extinction coefficients:
> >
> > As there are no Trp, Tyr or Cys in the region considered, your protein
> > should not be visible by UV spectrophotometry.
> > -----------
> >
> > There's a reason for it - while some absorption at 280 will indeed be
> > present, it won't be characteristic, and it would be overwhelmed by
> > non-specific absorption of crud.
> >
> > Otherwise, way to go Emir :)
> >
> > A.G.E.
> > "Emir Khatipov" <khatipovNO at NOuchicago.edu> wrote in message
> > news:pyul8.311$s4.23478 at news.uchicago.edu...
> > > Use Protparam tool at www.expasy.ch/tools to calculate molar
extinction
> > > coefficient of your peptide and use it for your spectrophotometric
> > > measurements. Peptides absorb light at 280nm even if they don't
contain
> > any
> > > aromatic residues. I hope it is needless to say that you should use
same
> > 10%
> > > DMSO in PBS as a reference. If you believe there are impurities in
your
> > > samples, I would suggest determining absorption spectra of your
peptide
> > > solutions at ~200-350nm (or at least 250-330nm). If your sample is
pure,
> > you
> > > should see a nice peak at around 276-280nm. Be aware that peptide bond
> > > absorbs at 214nm, but there might be a lot of interference from
> impurities
> > > you may have in your sample.
> > >
> > > Emir
> > >
> > >
> > > "JB" <j.bukczynskiDELETE_ME at utoronto.ca> wrote in message
> > > news:b87a9ugslasbigp8lm4dhkqc4p0soe0se7 at 4ax.com...
> > > > Hi,
> > > >
> > > > Does anyone know how to measure peptide concentration using a
> > > > spectrophotometer? I have it dissolved in 10% DMSO in PBS. Any help
> > > > would be appreciated. Thanks,
> > > >
> > > > Jacob
> > >
> > >
> >
> >
>
>





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