Recombinant Human SENP1, CF

Name: Recombinant Human SENP1, CF
Brand: R&D Systems
SKU: 6587-SP

Catalog #: 6587-SP Datasheet / COA / SDS

Newer Version Available: E-700

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Discontinued Product

6587-SP has been discontinued and is replaced by E-700.

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Recombinant Human SENP1, CF Summary

Product Specifications

Purity

>95%, by SDS-PAGE under reducing conditions and visualized by Colloidal Coomassie® Blue stain at 5 μg per lane

Endotoxin Level

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

Activity

Measured by its ability to proteolytically process pro-SUMO1. <10 ng of recombinant human SENP1 is required to cleave 50% of 1 μg of pro-SUMO1 or <25 ng of recombinant human SENP1 is required to cleave 95% of 1 μg of pro-SUMO1, as measured under the described conditions.

Source

E. coli-derived human SENP1 protein
Glu419-Leu644, with an N-terminal Met and 6-His tag.
Accession # Q9P0U3

Accession #

Q9P0U3

N-terminal Sequence
Analysis

Met

Predicted Molecular Mass

28 kDa

SDS-PAGE

27-28 kDa, reducing conditions

Product Datasheets

6587-SP

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.

6587-SP

Formulation	Supplied as a 0.2 μm filtered solution in Tris, NaCl, Glycerol, Brij-35 and DTT.
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

Assay Buffer: 50 mM Tris, 5 mM DTT, pH 8.5
Recombinant Human (rhSENP1) (Catalog # 6587-SP)
Recombinant Human SUMO1 (rhSUMO1) (Catalog # 3289-SU)
SDS-PAGE and/or Western blot
Densitometer (BioRad GS-800 or equivalent)

Dilute rhSUMO1 to 100 μg/mL in Assay Buffer.
Prepare a curve of rhSENP1 in Assay Buffer. Make the following serial dilutions: 4, 2, 1, 0.5, 0.25, 0.125, and 0.0625 µg/mL.
Combine 10 µL of each dilution with 10 µL of the 100 µg/mL rhSUMO1. Include an enzyme control containing Assay Buffer in place of rhSENP1.
Incubate the reaction mixtures and control at 37 °C for 20 minutes.
Combine 20 µL of reactions (and controls) with 20 µL reducing SDS-PAGE sample buffer.
Analyze the cleavage by SDS-PAGE (load 40 µL per lane) followed by protein staining and/or Western blot.
Determine the % of full length cleaved by densitometry and plot vs. rhSENP1 concentration with 4-PL fitting.
Activity Calculation:

% Cleavage = [1 -	% full-length rhSUMO1 (reaction)	] x 100%
	% full-length rhSUMO1 (control)

Per Well:

rhSUMO1: 1 µg
rhSENP1: 40, 20, 10, 5, 2.5, 1.25, 0.625, and 0 ng

Reconstitution Calculator

Background: SENP1

The small ubiquitin-like modifier (SUMO) is a member of ubiquitin-like protein family. SUMO modification of the target proteins is a reversible process that regulates many cellular processes including transcription regulation, nuclear localization, centromere segregation and signal transduction (1). Sentrin-specific proteases (SENPs) are a group of cysteine-type peptidases that catalyze two essential functions in the SUMO pathways: processing of full-length SUMOs to their mature forms and deconjugation of SUMOs from SUMOlyated proteins (2). The seven mammalian SENPs share a conserved C-terminal catalytic domain while the N-terminal domains have no significant similarity. Human SENP-1 has broad specificity for the three mammalian SUMOs (3). It is found in the cytoplasm and nucleus depending on cell type, and is expressed in testis, thymus, pancreas, spleen, liver, ovary and small intestine. It is thought that localization of SENP-1 is vital for the regulation for SUMOylation status of target proteins. The recombinant human SENP-1 represents the catalytic domain, which has been shown to be sufficient for SENP-1 activity and substrate specificity (3, 4).

References

Johnson, E.S. (2004) Annu. Rev. Biochem. 73:355.
Drag, M. and G.S. Salvesen (2008) IUBMB Life. 60:734.
Mikolajczyk, J. et al. (2007) J. Biol. Chem. 282:26217.
Xu, Z. and S.W.N. Au (2005) Biochem. J. 386:325.

Long Name

SUMO-Specific Peptidase 1

Entrez Gene IDs

29843 (Human); 223870 (Mouse); 300193 (Rat)

Alternate Names

EC 3.4.22; EC 3.4.22.-; SENP1; Sentrin/SUMO-specific protease SENP1; sentrin-specific protease 1; SUMO1/sentrin specific peptidase 1; SUMO1/sentrin specific protease 1; SuPr-2