10 Tips and Tricks for Protein DNA Crystallization
Pursuing the crystallization of protein-DNA complexes can wreck your publication rate. I have seen a number of colleagues dedicate years attempting to crystallize protein and DNA complexes, only to be unsuccessful in the end.
Here are a number of tips and tricks I hope you find helpful:
1) Adjust DNA parameters before protein
DNA plays a critical role in crystallization since it is often the source of crystal contacts.
2) Start with 10-11 base pairs of DNA
Increase 1-2 bases at a time, usually stopping at 21 bp. The idea is that a helix forms every 10.5 base pairs and forms a pseudo-continuous helix.
3) Sticky ends
The sticky ends do not have to be complementary. Blunt ends can work, but this would not be one of the first variables to change.
4) Screening
Expect all the normal fun of protein screening. I would favor PEG or MPD in screens (Natrix) rather than high salt screens. The high salt can disrupt the charged interaction between the protein and DNA.
5) Get your DNA from IDT
We have also ordered through GeneArt
6) Run a gel
If you are unsure if you have protein-DNA complex formation
7) Don’t worry about purification
This is assuming your DNA is from a commercial resource (it has already been purified). If you are running out of options, a quick run over a HPLC may help.
8 ) Concentrate your protein-DNA before crystallization trials
If precipitation is occurring, consider running DLS to help determine at what concentrations precipitation is occurring
9) Use a ratio for protein to DNA of 1:1.2-5
10) Cryoprotect your Crystals
If your crystallization condition does not contain a cryoprotectant then try a second pass through one of your favorites, like glycerol or LV Oil.