Try adjusting pH or salt content. Arginine in positively charged so pH could be involved. Other thought would be to express with GST tag and purify with appropriate resin?

You can play with two parameters. Try to increase the working pH, if the protein does not aggregate. That might increase the binding efficiency. Decrease the flow-rate to make the binding efficient. Do not use more resin that might increase non-specific binding to the resin. If either of this work then play with NaCl conn as well

Is your arginine rich protein tagged with 6xHis? Where did you tagged 6x His on your protein, that is, at N-terminus or at C-terminus? In case of C-terminal 6xHis tagging, I have similar experience with you and solved the problem by tagging 6xHis at N-terminal.

Have you run a western to make sure that your his-tagged protein is really his-tagged?

You might try a different resin- sometimes I have better luck with Cobalt-based resin than Nickel-based resin. For example, http://www.clontech.com/US/Products/Protein_Expression_and_Purification/His-Tagged_Protein_Purification/Cobalt_Resin-Batch

I also agree that you should play with pH, incubation time, and/or try switching the His-tag to the other terminus. You could also try co-expressing your protein of interest with a known binding partner. This would stabilize the structure while binding, and then you could use a high salt wash to remove it. This method worked for me with a few hard-to-purify proteins.

Here are more specifics:

Lysis buffer is 20mM CAPS 10.5; 50mM NaCl; 5mM B-ME; 1mM PMSF

Gravity flow column with Ni-NTA from Promega. Recently purchased.

Lowering the pH causes the protein to crash out.

Increasing the NaCl concentration to 0.5M does not alter the described Ni-NTA binding behavior.

There is a fusion (SUMO) protein at the N-terminus with a His-Tag at the N-terminus of SUMO. This is routinely done.

Moving His-tag to C-terminus may be a possible option, but I'm concerned that this may have an deleterious effect on the biological activity of the protein, as there is published evidence suggesting this.

Why CAPS buffer? This is a secondary amine, which may interfere with His-tag binding. Perhaps try a simple sodium phosphate buffer.

CAPS has worked for a number of other His-Tag proteins I've worked with; however, I'll try phosphate as a replacement for the next purification.

The issue with poor binding may be because of the gravity flow method not giving the protein enough time to bind. I would incubate the lysate with the media at 5oC with gentle rocking (to keep the media suspended) for 1-2 hrs and then pour the media/lysate into the column and proceed as normal.

I just had a similar problem with the Ni-NTA column. Keep in mind the pH of the buffer changes rapidly if you are using UREA. Need to measure pH of buffer moments before you use it!

I have tried anion and cation exchange. Cation works to a certain extent, but Ni-NTA purification is a lot easier if I can get it to work.

Have you contacted Promega Tech support about the compatibility of CAPS buffer and Ni-NTA media? CAPS is not listed as being compatible with Ni-NTA media, see attached file. I saw you have used CAPS before and it worked, are you preparing fresh stocks of CAPS buffer for each purification? And are you using new media/resin for each purification?

If it is possible, carry out the purification at pH 7-8. In case, lower the flow and play with salt content. Is the protein really soluble at pH 7-8?

I have not contacted Promega. The CAPS is 2 months old, but kept in the dark as I think it's light sensitive (?). I've used CAPS that is close to 1 year old with no problems. Also, CAPS is supposed to have very low metal binding capacity. It was used as a buffer to study metal binding enzymes. There are two articles in Methods Enzymology that cover this (Good & Izawa 1972, Blanchard 1984).

I'll try sodium phosphate at this pH to see what happens. I'm a little concerned that the phosphate may cause other problems, but no telling unless I try.

Lowering the pH results in precipitation. I've tried a lot of additives to inhibit precipitation, but no luck.

Just on a side note:

Imidazole buffers perfectly fine in the pH range used for most Ni-NTA purifications.

Just add everything you think you need (salts etc.), adjust your imidazole concentration, pH all buffers to the desired value, done.

Oh, and for those Arginines, take the NaCl out (all of it) and use ~300 mM K-Glutamate at pH ~7.5 as salt. That should be more gentle to your protein than Chloride.

You might want to try running your sample over a different ion exchange column first to get rid of other proteins that may be interfering with your protein of interest binding to Ni-NTA. Then run the pass through containing your protein of interest over the Ni-NTA.

The K-Glutamate is an interesting idea. I'll give it a try. Is it easy to dialyze out?

Should be no problem. But if it helps, why remove it? You can also try Ammonium Acetate or something like that as salt.