Protein convertase subtilisin/kexin 9 (PCSK9) possesses a signal peptide, a prosegment, a catalytic domain, a hinge region, and a C-terminal Cys-His-rich domain. This protein is primarily synthesized and secreted by hepatocytes. Besides the liver, it is also expressed in the small intestine, kidney, and brain, and it is present in plasma. Upon translocation to the endoplasmic reticulum, the prosegment of PCSK9 is autocatalytically cleaved and secreted as a stable, enzymatically inactive, non-covalent complex. Elevated low density lipoprotein-cholesterol (LDLc) levels are a major risk factor for cardiovascular disease and atherosclerosis. PCSK9 is a regulator of LDLc levels through the binding of the LDL-Receptor (LDLR) which subsequently leads to its degradation. PCSK9 null mice fed with a high fat diet demonstrated a four-fold decrease in cholesterol accumulation, which stresses the important role played by PCSK9 in the development of cardiovascular diseases. PCSK9 also binds to other LDLR family members such as very low density lipoprotein receptor (VLDLR), apolipoprotein E receptor (LRP1/APOER), and apolipoprotein receptor 2 (LRP8/APOER2), which leads to their degradation in the intracellular acidic compartments. Like the LDLR, gene expression of PCSK9 is positively regulated by sterol-regulatory-element-binding protein-2 (SREBP-2), a transcription factor that is activated in response to cellular cholesterol depletion.
Mouse PCSK9, (amino acids Gln35-Gln694, Accession# NP_705793), was expressed with a C-terminal His tag and a linker sequence in a 293E cells.
Molecular Mass
The 709 amino acid recombinant protein has a predicted molecular mass of approximately 76.6 kD. The mature and pro-domain recombinant proteins migrate at 70 kD and 17 kD in DTT-reducing conditions and at 63 kD and 17 kD in non-reducing conditions, respectively, by SDS-PAGE. The predicted N-terminal amino acid is Gln.
Purity
>95%, as determined by Coomassie stained SDS-PAGE
Formulation
Lyophilized in sterile-filtered PBS, pH 7.2, containing 1% BSA, 0.09% sodium azide, and protease inhibitors.
Concentration
Lot-specific (please contact technical support for mass/vial, or use our Lookup tool if you have a lot number.)
Storage & Handling
Upon receipt, store unopened vials between 2°C and 8°C immediately and use within 12 months from date of receipt. Prior to use, reconstitute the lyophilized powder with 0.2 ml of PBS containing a carrier protein (e.g., 1% BSA, protease free), pH 7.4. Re-cap vial, vortex. Allow the reconstituted standard to sit at room temperature for 15 minutes, vortex again to mix completely. The reconstituted standard stock solution can be aliquoted into polypropylene vials and stored at -70°C for up to one month. Do not re-use diluted standards. Use a manual defrost freezer and avoid repeated freeze thaw cycles.
Each lot of this protein is quality control tested by ELISA. For use as an ELISA standard, a standard curve comprised of doubling dilutions from 4000 pg/mL to 62.5 pg/mL is suggested. It is recommended that the reagent be titrated for optimal performance.
Application Notes
This PCSK9 protein is useful as a standard for a mouse PCSK9 sandwich ELISA, using unlabeled M114G10 antibody as capture and biotinylated Poly5251 antibody as detection.
