Recombinant Mouse Prss28 Protein, CF

Catalog #: 4808-SE Datasheet / COA / SDS

Catalog #	Availability	Size / Price	Qty
4808-SE-010

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Recombinant Mouse Prss28 Protein, CF Summary

Product Specifications

Purity

>90%, by SDS-PAGE under reducing conditions and visualized by silver stain

Endotoxin Level

<1.0 EU per 1 μg of the protein by the LAL method.

Activity

Measured by its ability to cleave a peptide substrate, Suc-VPF-SBzl after activation by Thermolysin. The specific activity is >15,000 pmol/min/µg, as measured under the described conditions.

Source

Mouse myeloma cell line, NS0-derived mouse PRSS28 protein
Ala19-Gln274, with a C-terminal 10-His tag
Accession # Q924N9

Accession #

Q924N9

N-terminal Sequence
Analysis

Ala19

Structure / Form

Pro form

Predicted Molecular Mass

30 kDa

SDS-PAGE

34 kDa, reducing conditions

Product Datasheets

4808-SE

Product Datasheet

COA

SDS

Carrier Free

What does CF mean?

CF stands for Carrier Free (CF). We typically add Bovine Serum Albumin (BSA) as a carrier protein to our recombinant proteins. Adding a carrier protein enhances protein stability, increases shelf-life, and allows the recombinant protein to be stored at a more dilute concentration. The carrier free version does not contain BSA.

What formulation is right for me?

In general, we advise purchasing the recombinant protein with BSA for use in cell or tissue culture, or as an ELISA standard. In contrast, the carrier free protein is recommended for applications, in which the presence of BSA could interfere.

4808-SE

Formulation	Supplied as a 0.2 μm filtered solution in Tris and NaCl.
Shipping	The product is shipped with dry ice or equivalent. Upon receipt, store it immediately at the temperature recommended below.
Stability & Storage:	Use a manual defrost freezer and avoid repeated freeze-thaw cycles. 6 months from date of receipt, -70 °C as supplied. 3 months, -70 °C under sterile conditions after opening.

Assay Procedure

Materials

Activation Buffer: 50 mM Tris, 150 mM NaCl, 10 mM CaCl₂, 0.05% Brij-35, pH 7.5 (TCNB)
Assay Buffer: 50 mM Tris, 150 mM NaCl, pH 8.0
Recombinant Mouse PRSS28 (rmPRSS28) (Catalog # 4808-SE)
Bacterial Thermolysin (Catalog # 3097-ZN)
1,10-Phenanthroline (Sigma, Catalog # 320056), 600 mM stock in DMSO
Substrate: SUC-Val-Pro-Phe-SBZL (Bachem, Catalog # M-1885), 10 mM stock in DMSO
5,5’-dithio-bis(2-nitrobenzoic acid) (DTNB) (Sigma, Catalog # D-8130), 10 mM stock in DMSO
96 well clear plate (Costar, Catalog # 92592)
Plate Reader (Model: SpectraMax Plus by Molecular Devices) or equivalent

Dilute rmPRSS28 to 100 µg/mL with 0.1 µg/mL Thermolysin in Activation Buffer.
Incubate at 37 °C for 60 minutes.
Stop Thermolysin activity by adding 1,10-Phenanthroline to 10 mM.
Incubate at room temperature for 15 minutes.
Dilute active rmPRSS8 to 0.05 µg/mL in Assay Buffer.
Dilute Substrate to 200 µM with 200 µM DTNB in Assay Buffer.
Load 50 µL of the 0.05 µg/mL rmPRSS28 into a clear microplate. Include a Substrate Blank containing 50 µL of Assay Buffer.
Start the reaction by adding 50 µL of the Substrate Mixture into wells.
Read in kinetic mode for 5 minutes at an absorbance of 405 nm.
Calculate specific activity using the following formula:

Specific Activity (pmol/min/µg) =	Adjusted V_max* (OD/min) x well volume (L) x 10¹² pmol/mol
	ext. coeff (M^-1cm^-1) x path corr.* (cm) x amount of enzyme (µg)

     *Adjusted for Substrate Blank
      **Using the extinction coefficient 13260 M^-1cm^-1
      ***Using the path correction 0.320 cm
     Note: the output of many spectrophotometers is in mOD. Per Well:

rmPRSS8: 2.5 ng
Substrate: 100 µM
DTNB: 100 µM

Reconstitution Calculator

Background: PRSS28

Mouse Prss28, also known as Implantation Serine Protease 1 (ISP1), belongs to the S1 serine proteinase family with a conserved Histidine-Aspartic Acid-Serine catalytic triad (1). It closely resembles the mouse mast cell tryptases, specifically MCP-6 and MCP-7 (1). It is co-expressed with ISP2/Prss29 in pre-implantation embryos and uterus (2). It is involved with embryo hatching and its activity is important for successful implantation (2).

References