Reducing DNA content during protein purification
We have just published our poster for “Reducing DNA content during protein purification with an easily removable nuclease” to the website. This poster was presented at PepTalk, January 19-23 2015 in San Diego, USA. Below you can read the introduction:
When purifying DNA-modifying enzymes, an essentially DNA-free product is desirable as endogenous DNA from production may contaminate down-stream applications.
While nuclease treatment of the crude cell-lysate is a common step during protein purification, its primary function is viscosity reduction for easier sample handling rather than complete DNA removal. More efficient enzymatic DNA removal could be achieved if applied to a partially purified protein sample, such as the His-trap eluate, where the majority of impurities have been removed and the volume is reduced; however the salt concentrations used in elution protocols are incompatible with the salinity optima of most commercially available nucleases. Furthermore, treatment of semi-pure protein may cause residual nuclease contamination in the purified protein which will interfere with
nucleic-acid based applications.
ArcticZymes has developed a Heat-Labile Salt Active Nuclease (HL-SAN) that is compatible with treatment of eluates: it is active at up to 1M NaCl, tolerates imidazole and can be separated from most proteins by cation exchange or inactivated by reducing agents. Here we present efficient DNA removal from recombinantly-produced T4 DNA ligase by direct treatment of the His-trap eluate with HL-SAN, and demonstrate that the final, active T4 ligase is free of contaminating HL-SAN activity.
You can view the poster here (PDF) or learn more about HL-SAN over here