HL-SAN, ArcticZymes Technologies ASA, Norway
Removal of nucleic acids may improve workflows during protein purification, both for laboratory sample preparation and industrial bioprocessing. HL-SAN is a nonspecific endonuclease with optimum activity at high salt concentrations. HL-SAN is active in a variety of buffers and can be easily inactivated by treatment with a reducing agent. These features make HL-SAN particularly useful in the purification of proteins and removal of DNA and RNA from molecular biology reagents.
Nucleic acids, and especially genomic DNA, often pose a problem in purification of DNA-binding proteins as they interfere with purification, downstream analysis or applications.
As most nucleases are inhibited by high concentrations of salt, removal of DNA is difficult, since NaCl is used to dissociate DNA from proteins.
The high salt-tolerance and easy removal makes HL-SAN beneficial to use in protein purification schemes, especially in combination with IMAC (Immobilized metal affinity chromatography) media used for purification of poly-His-tagged proteins both in conventional chromatography or high-throughput settings.
Nonspecific endonuclease
Active at high salt conditions
Easily
inactivated
Properties
| Source | Recombinantly produced in Pichia pastoris. |
| Activity | HL-SAN is highly active in the temperature range 10–50°C. Optimal NaCl-concentration for activity is 0.5 M, working range is 0.25–1 M. Mg2+ (>1 mM) is required for activity. Working pH range is 7.5–9.5, optimal pH is 9.0. |
| Specific activity | ≥ 175 000 Units/mg |
| Unit definition | One Unit is defined as an increase in absorbance at 260 nm of 1 A in 30 minutes at 37°C, using 50 μg/ml calf thymus DNA (D-1501, Sigma) in a buffer consisting of 25 mM Tris-HCl, pH 8.5 (25°C), 5 mM MgCl2, 500 mM NaCl. |
Tolerance to typical lysis buffer additives
| Imidazole | 20% activity at 350 mM Imidazole |
| Glycerol | 20% activity at 35% glycerol |
| Triton X-100 | No reduced activity in Triton X-100 (tested up to 15%) |
| SDS | Not recommended |
| Urea | Not recommended |
Publications
HL-SAN Applications
- Nanopore metagenomics enables rapid clinical diagnosis of bacterial lower respiratory infection.
Charalampous T, Kay GL, Richardson H, Aydin A, Baldan R., Jeanes C, Rae D, Grundy S, Turner DJ, Wain J, Leggett RM, Livermore DM, O’Grady J.
Nature Biotechnology. 2019; 37: 783–792. - A Structured Workflow for Mapping Human Sin3 Histone Deacetylase Complex Interactions Using Halo-MudPIT Affinity-Purification Mass Spectrometry.
Banks CAS, Thornton JL, Eubanks CG, Adams MK, Miah S, Boanca G, Liu X, Katt M, Parmely T, Florens LA, Washburn MP.
Molecular & Cellular Proteomics. 2018; 17 (7): 1432-1447. - Biochemical characterization of ParI, an orphan C5-DNA methyltransferase from Psychrobacter arcticus 273-4.
Grgic M, Williamson A, Kjaereng Bjerga GE, Altermark B, Leiros I.
Protein Expr Purif. 2018; 150: 100-108. - Biochemical Characterization of a Family 15 Carbohydrate Esterase from a Bacterial Marine Arctic Metagenome.
De Santi C, Willassen NP, Williamson A.
PLoS ONE. 2016; 11(7): e0159345. - Recombinant expression and purification of an ATP-dependent DNA ligase from Aliivibrio salmonicida.
Williamson A, Pedersen H.
Protein Expr Purif. 2014; 97: 29-36.
