Human Prostasin/PRSS8 HEK293 Overexpression Lysate

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Human Prostasin/PRSS8 HEK293 Overexpression Lysate: Product Information

Product Description
This Human Prostasin/PRSS8 overexpression lysate was created in HEK293 Cells and intented for use as a Western blot (WB) positive control. Purification of Prostasin/PRSS8 protein (Cat: 10287-H08H) from the overexpression lysate was verified.
Expression Host
HEK293 Cells
Species
Human
Sequence Information
A DNA sequence encoding the mature form of human Prss8 (NP_002764.1) (Ala 30-Arg 322) was fused with a signal peptide at the N-terminus and a polyhistidine-tag at the C-terminus.
Molecule Mass
The mature recombinant human Prss8 consists of 304 amino acids and predicts a molecular mass of 32.8 kDa. By SDS-PAGE under reducing conditions, the apparent molecular mass of rhPrss8 is approximately 40 kDa due to glycosylation.

Human Prostasin/PRSS8 HEK293 Overexpression Lysate: Usage Guide

Preparation Method
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.
Lysis Buffer
Modified RIPA Lysis Buffer: 50 mM Tris-HCl pH 7.4, 150 mM NaCl, 1mM EDTA, 1% Triton X-100, 0.1% SDS, 1% Sodium deoxycholate, 1mM PMSF.
Recommend Usage
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.
Sample Buffer
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.
Application
Western Blot (WB)
Optimal dilutions/concentrations should be determined by the end user.

Human Prostasin/PRSS8 HEK293 Overexpression Lysate: Alternative Names

Human CAP1 Overexpression Lysate; Human PROSTASIN Overexpression Lysate

Prostasin/PRSS8 Background Information

Prostasin (Prss8), also known as channel activating protease 1 (CAP1), is a trypsinlike serine peptidase, and plays important roles in epithelial physiology. It is originally purified as an active, soluble enzyme from human seminal fluid and is highly expressed in prostate, lung, kidney, salivary gland and pancreas. Prostasin is expressed as a glycosyl-phosphatidylinositol (GPI)-anchored membrane protein in prostate epithelial cells, and also exists as a secreted proteolytic enzyme possibly via tryptic cleavage of its COOH-terminal hydrophobic domain. Prostasin is found to activate the epithelial sodium channel (ENaC) which is tightly regulated and is critical for maintaining salt and fluid balance in the lung and kidney in both normal and pathological conditions. Accordingly, prostasin has been proposed as a target for therapeutic inhibition in cystic fibrosis. In addition, prostasin inhibits prostate and breast cancer cell invasion in vitro, suggesting a functional role as a suppressor of tumor invasion, as well as a regulator of gene expression during inflammation.
Full Name
protease, serine, 8
References
  • Yu J.X., et al.,(1995), Molecular cloning, tissue-specific expression, and cellular localization of human prostasin mRNA. J. Biol. Chem. 270:13483-13489.
  • Ota T., et al., (2004), Complete sequencing and characterization of 21,243 full-length human cDNAs.Nat. Genet. 36:40-45.
  • Martin J., et al.,(2004), The sequence and analysis of duplication-rich human chromosome 16.Nature 432:988-994.
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