Human ORP150 HEK293 Overexpression Lysate: Product Information
This Human ORP150 overexpression lysate was created in HEK293 Cells and intented for use as a Western blot (WB) positive control. Purification of ORP150 protein (Cat: 11342-H08H) from the overexpression lysate was verified.
A DNA sequence encoding the C-terminal segment of human HSP12A (NP_001124463.1) (Met 695-Leu 994) was expressed, fused with a polyhistidine tag at the C-terminus and a signal peptide at the N-terminus.
The recombinant human HSP12A consists of 311 amino acids and predictes a molecular mass of 35.2 kDa. In SDS-PAGE under reducing conditions, the apparent molecular mass of rh HSP12A is approximately 55-65 kDa due to glycosylation.
Human ORP150 HEK293 Overexpression Lysate: Usage Guide
Cell lysate was prepared by homogenization of the over-expressed cells in ice-cold modified RIPA Lysis Buffer with cocktail of protease inhibitors (Sigma). Cell debris was removed by centrifugation. Protein concentration was determined by Bradford assay (Bio-Rad protein assay, Microplate Standard assay). The cell lysate was boiled for 5 min in 1 x SDS loading buffer (50 mM Tris-HCl pH 6.8, 12.5% glycerol, 1% sodium dodecylsulfate, 0.01% bromophenol blue) containing 5% b-mercaptoethanol, and lyophilized.
1. Centrifuge the tube for a few seconds and ensure the pellet at the bottom of the tube.
2. Re-dissolve the pellet using 200μL pure water and boil for 2-5 min.
1 X Sample Buffer (1 X modified RIPA buffer+1 X SDS loading buffer).
Stability & Storage
Store at 4℃ for up to twelve months from date of receipt. After re-dissolution, aliquot and store at -80℃ for up to twelve months. Avoid repeated freeze-thaw cycles.
Western Blot (WB) Optimal dilutions/concentrations should be determined by the end user.
Human ORP150 HEK293 Overexpression Lysate: Alternative Names
Human GRP-170 Overexpression Lysate; Human Grp170 Overexpression Lysate; Human HSP12A Overexpression Lysate; Human ORP-150 Overexpression Lysate; Human ORP150 Overexpression Lysate
ORP150 Background Information
Hypoxia up-regulated protein 1, also known as 15 kDa oxygen-regulated protein, 17 kDa glucose-regulated protein, ORP-15, GRP-17 and HYOU1, is a member of theheat shock protein 7 family. Seven members from four different heat shock protein (HSP) families were identified including HYOU1 (ORP15), HSPC1 (HSP86), HSPA5 (Bip), HSPD1 (HSP6), and several isoforms of the two testis-specific HSP7 chaperones HSPA2 and HSPA1L. HYOU1 is highly expressed in tissues that contain well-developed endoplasmic reticulum and synthesize large amounts of secretory proteins. It is highly expressed in liver and pancreas. HYOU1 is also expressed in macrophages within aortic atherosclerotic plaques, and in breast cancers. HYOU1 has a pivotal role in cytoprotective cellular mechanisms triggered by oxygen deprivation. It may play a role as a molecular chaperone and participate in protein folding. Suppression of HYOU1 is associated with accelerated apoptosis. It is suggested to have an important cytoprotective role in hypoxia-induced cellular perturbation. This protein has been shown to be up-regulated in tumors, especially in breast tumors, and thus it is associated with tumor invasiveness.
hypoxia up-regulated 1
Ikeda J., et al.,(1997), Cloning and expression of cDNA encoding the human 150 kDa oxygen-regulated protein, ORP150. Biochem. Biophys. Res. Commun. 230:94-99.
Kaneda S., et al., (2000), Production of three distinct mRNAs of 150 kDa oxygen-regulated protein (ORP150) by alternative promoters: preferential induction of one species under stress conditions.J. Biochem. 128:529-538.
Takeuchi S., (2006), Molecular cloning, sequence, function and structural basis of human heart 150 kDa oxygen-regulated protein, an ER chaperone.Protein J. 25:517-528.
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