Recombinant Shrimp Alkaline Phosphatase (rSAP)
The gold standard for thermolabile alkaline phosphatase has been improved further. A recombinant version of SAP has been developed and this version adds further advantages to the product. The current recombinant version is produced in Pichia pastoris, which ensures a recombinant enzyme that is fully active and correctly expressed, at consistently high purity in large batches.
Main advantages with recombinant SAP
- Identical catalytic properties with the native version
- High purity that meets the highest standards
- Available in large batches; less need for batch qualification
- Recombinant production ensures minimal batch-to-batch variation
- Excellent stability for months at 4°C and room temperature
- Available in high concentrations
- Ideally suited for high throughput robotic systems
- The same reliable performance as the well-proven native SAP
Source
Recombinant SAP is produced from fermentation of a Pichia pastoris clone carrying several copies of a cDNA originally isolated from Arctic Shrimp, coding for alkaline phosphatase. Fusion to a secretion signal has facilitated secretion of the expression product into the fermentate. Correct signal peptide cleavage has been verified by N-terminal sequencing of the recombinant product. Recombinant SAP is otherwise identical in amino acid sequence to the native enzyme, no modifications (tags or similar) have been added.
Specifications
Unit definition: One Unit will convert 1 µmol of p-nitrophenyl phosphate per minute to nitrophenol and phosphate at 37ºC and pH 10.4 in 0.1 M glycine buffer, 1 mM each of ZnCl2 and MgCl2.
This Unit definition makes 1 Unit of SAP equivalent to 5 to 40 Units of Antarctic Phosphatase (New England Biolabs), 1.3 Units APex phosphatase or 35 Units NTPhos phosphatase (Epicentre Biotechnologies). SAP is therefore the most processive alternative.
Specific activity: >2 000 Units/mg.
Purity: Purified to apparent homogeneity by SDS-PAGE. DNase, RNase or protease activities not detected.
Concentration: Minimum 10 000 Units/ml, available up to 30 000 Units/ml.
Batch sizes: 25 – 35 million Units / batch.
Properties
Stability: Stable at -20ºC in storage buffer (25 mM Tris-HCl pH 7.6, 5 mM MgCl2, glycerol 50%). At room temperature, >95% activity remains after 90 days.
Still, the enzyme is completely inactivated after 5 minutes at 65°C. At 75°C, rSAP is completely inactivated after only 1 minute. In a standard thermocycler, the process of heating from 37 to 95 and back to 37°C is sufficient to completely inactivate SAP.
Availability
The product is now available directly from ArcticZymes, both as bulk product for OEM, and in convenient end-user packs through our web shop. The product is also available through Affymetrix (USB).
Intellectual property
Recombinant SAP is covered by the following patents: US 7,323,325, EP 1,326,890, JP 4,191,479, AU 9,398,701 and related applications in other countries either granted or pending.