Arg319-Phe541. N-glycosylation site at Asn (N) 331 was mutated to Gln (Q).
Expression System
HEK293 cells
Tag
C-terminal His-tag
Purification
His-tag affinity purification by immobilized metal ion affinity chromatography (IMAC)
Purity
>95 %
Purity Determined By
SDS-PAGE under reducing conditions and visualized by Coomassie blue staining
Molecular Weight (kDa)
Recombinant protein product has a calculated molecular mass of 25 kDa. Due to the abundant glycosylation, it migrates as approximately 30 kDa protein bands in SDS-PAGE under DTT, beta-mercaptoethanol reducing conditions. See deglycosylation analysis image below.
Format
Liquid
Formulation
Supplied as a 0.2 µm filtered solution in PBS (pH 7.4)
Concentration (lot specific)
Lot specific (see the label on the vial), determined by BCA protein assay
Figure 1. Deglycosylation of purified recombinant proteins. Purified proteins were untreated (Lane 2) or treated with Protein Deglycosylation Kit under native (Lane 3) or reducing (Lane 4) conditions. Deglycosylation treatment resulted in a mobility shift of the protein to produce one major band at the expected size, thus indicating that the untreated recombinant protein (Lane 2) was glycosylated. Lane 1: Protein standard ladder (kDa) Lane 2: Untreated protein under reducing conditions Lane 3: Treated protein with deglycosylation enzymes under native conditions Lane 4: Treated protein with deglycosylation enzymes under reducing conditions.
Binding Function
Product binding ability was measured by ELISA. The immobilized recombinant human ACE2 protein (Catalog #230-30165, coated at 0.5 µg/ml, 100 µl/well) was incubated with the serial diluted SARS CoV-2 S1 RBD N331 mutant protein (Catalog # 230-30157). The bound mutant protein was detected by mouse anti-S1 RBD monoclonal antibody using ELISA. The calculated EC50 is 138.1 ng/ml.
Stability & Storage
Shipping
Ice packs
Storage/Stability
Upon arrival, the protein may be stored for 2 weeks at 4 °C. For long term storage, it is recommended to store at -20 °C or -80 °C in appropriate aliquots. Avoid repeated freeze-thaw cycles.
