Human HRG/HPRG HEK293 Overexpression Lysate: Product Information
This Human HRG/HPRG overexpression lysate was created in HEK293 Cells and intented for use as a Western blot (WB) positive control. Purification of HRG/HPRG protein (Cat: 10836-H08H) from the overexpression lysate was verified.
A DNA sequence encoding the human HPRG (NP_000403.1) (Met 1-Lys 525) with a C-terminal polyhistidine tag was expressed.
The secreted recombinant human HPRG comprises 518 amino acids with a predicted molecular mass of 59 kDa. As a result of glycosylation, rhHPRG migrates as an approximately 75-80 kDa band in SDS-PAGE under reducing conditions.
Human HRG/HPRG 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 HRG/HPRG HEK293 Overexpression Lysate: Alternative Names
Human HPRG Overexpression Lysate; Human HRGP Overexpression Lysate; Human THPH11 Overexpression Lysate
HRG/HPRG Background Information
Histidine-rich glycoprotein, also known as HRG and HPRG, is a glycoprotein located in plasma and platelets, and contains an unusually large amount of histidine and proline. In human, five distinct domains are recognized in the mature HPRG molecule. There are two N-terminal cystatin-like modules (aa 19 - 254) and one His-Pro-rich region (aa 35 - 497) that is flanked by two Pro-rich segments (aa 276 - 321 and 498 - 525). The His-Pro-rich region contains 1 tandem repeats with an HHPHG motif, and the N- and C-termini are linked by a disulfide bond. The specific functions of HRG remain unclear, but it is known that the protein binds heme, dyes and divalent metal ions. It inhibits rosette formation and interacts with heparin, thrombospondin and plasminogen. Two of the protein's effects, the inhibition of fibrinolysis and the reduction of inhibition of coagulation, indicate a potential prothrombotic effect. HPRG is evolutionarily, functionally and structurally related to cleaved high molecular weight kininogen (HKa), an anti-angiogenic polypeptide that stimulates apoptosis of proliferating endothelial cells through binding to cell-surface tropomyosin. The antiangiogenic activity of the multidomain plasma protein HPRG is localized to its histidine-proline-rich (H/P) domain and has recently been shown to be mediated, at least partially, through binding to cell-surface tropomyosin in fibroblast growth factor-2-activated endothelial cells.
Guan X, et al. (2004) Histidine-proline rich glycoprotein (HPRG) binds and transduces anti-angiogenic signals through cell surface tropomyosin on endothelial cells. Thromb Haemost. 92(2): 403-12.
Doate F, et al. (2004) Peptides derived from the histidine-proline domain of the histidine-proline-rich glycoprotein bind to tropomyosin and have antiangiogenic and antitumor activities. Cancer Res. 64(16): 5812-7.
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