In response to your second question, yes it is possible to separate
peptides and proteins that differ by a single charge. Some examples are to
be found in Section IV of "High Performance Liquid Chromatography of
Peptides and Proteins" edited by Colin T. Mant and Robert S. Hodges. Note
the chapters by Mant/Hodges, and Alpert.
Russ Lehrman
chris halkides <halkidesc@uncwil.edu>
06/23/98 12:13 PM
To: Recipients of ABRF List <abrf@aecom.yu.edu>
cc: (bcc: Russ Lehrman/NeXstar)
Subject: iex hplc of proteins
To All,
I am alkylating proteins in such a way that the product should
differ in charge from the starting material by 1-2 charges, depending on
pH. I have used RP HPLC to separate these species with success for the
smallest member of this family of proteins, but RP HPLC is less effective
for larger proteins, so far. I would like to try IEX HPLC, but I lack
experience with this technique. "Protein Purification" (Jansen and Ryden)
gives a nice example of this for a 73 residue protein, so clearly the
method has potential.
1. Is there a general principle about whether it is better to use
cation versus anion exchange hplc to separate protein species that differ
by 1 or two charges? I would have thought that one always chose the type
of charge on the ion-exchanger based on the overall charge of the protein,
but I have been told that this is not necessarily so, given that any
protein may have patches of positive and negative charge.
2. Can large proteins be separated if they differ by only one charge?
Are there references out there to work in which this has been done?
Thanks for any suggestions,
Chris Halkides
Christopher Halkides
Dept. of Chemistry, UNCW
601 S. College Road
Wilmington, NC 28403-3297