Recombinant Human Insulysin/IDE Protein, CF

Catalog #: 2496-ZN
7 Citations Datasheet / COA / SDS

Catalog #	Availability	Size / Price	Qty
2496-ZN-010

R&D Systems Recombinant Proteins and Enzymes

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Citations (7)

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Summary Product Datasheets Carrier Free Data Images Reconstitution Calculator Background Related Research Areas

Recombinant Human Insulysin/IDE Protein, CF Summary

Product Specifications

Purity

>95%, 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 the fluorogenic peptide substrate, Mca-RPPGFSAFK(Dnp)-OH (Catalog # ES005). The specific activity is >1,000 pmol/min/µg, as measured under the described conditions.

Source

Spodoptera frugiperda, Sf 21 (baculovirus)-derived human Insulysin/IDE protein
Met42-Leu1019, with an N-terminal Met and 7-His tag

Accession #

P14735

N-terminal Sequence
Analysis

Met

Predicted Molecular Mass

114 kDa

SDS-PAGE

105 kDa, reducing conditions

Product Datasheets

2496-ZN

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.

2496-ZN

Formulation	Supplied as a 0.2 μm filtered solution in Tris, NaCl, Brij-35 and Glycerol.
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, -20 to -70 °C as supplied. 3 months, -20 to -70 °C under sterile conditions after opening.

Assay Procedure

Materials

Assay Buffer: 50 mM Tris, 1 M NaCl pH 7.5
Recombinant Human Insulysin/IDE (rhInsulysin) (Catalog # 2496-ZN)
Substrate: MCA-Arg-Pro-Pro-Gly-Phe-Ser-Ala-Phe-Lys(DNP)-OH (Catalog # ES005), 2 mM stock in DMSO
F16 Black Maxisorp Plate (Nunc, Catalog # 475515)
Fluorescent Plate Reader (Model: SpectraMax Gemini EM by Molecular Devices) or equivalent

Dilute rhInsulysin to 0.2 µg/mL in Assay Buffer.
Dilute Substrate to 20 µM in Assay Buffer.
Load 50 µL of the 0.2 µg/mL rhInsulysin into a plate, and start the reaction by adding 50 µL of 20 µM Substrate. Include a Substrate Blank containing 50 µL Assay Buffer and 50 µL of 20 µM Substrate.
Read at excitation and emission wavelengths of 320 nm and 405 nm, respectively, in kinetic mode for 5 minutes.
Calculate specific activity:

Specific Activity (pmol/min/µg) =	Adjusted V_max* (RFU/min) x Conversion Factor** (pmol/RFU)
	amount of enzyme (µg)

*Adjusted for Substrate Blank

**Derived using calibration standard MCA-Pro-Leu-OH (Bachem, Catalog # M-1975).

Per Well:

rhInsulysin: 0.01 µg
Substrate: 10 µM

Reconstitution Calculator

Background: Insulysin/IDE

Insulysin, or insulin-degrading enzyme (IDE), is a zinc metallopeptidase of the inverzincin family. IDE is primarily located in the cytosol, but has been detected as a secreted enzyme and associated with the plasma membrane as well (1). The enzyme is expressed in many tissues, with the highest levels in liver, kidney, brain, and testis (2). IDE hydrolyzes a variety of regulatory peptides, including insulin, glucagon, atrial natriuretic factor, and transforming growth factor-alpha in vitro (1). In addition, IDE has been shown to degrade the amyloid beta (A beta ) peptide, which polymerizes into the plaques associated with Alzheimer's disease (3). Deficiencies in IDE activity may contribute to the pathogenesis of type 2 diabetes mellitus (DM2) and Alzheimer's disease. The IDE region of human chromosome 10q has been genetically linked to DM2 (4). When the IDE gene was specifically disrupted in mice, IDE -/- animals developed hyperinsulinemia and glucose intolerance, characteristics of DM2 (5). The IDE -/- mice were also shown to have a significant decrease in A beta degradation in the brain, resulting in increased cerebral accumulation of A beta peptide. This in vivo evidence is consistent with the hypotheses that IDE is important for the degradation of insulin in cells and for the clearance of A beta peptide in the brain.

References

Affholter, J. A. et al. (1988) Science 242:1415.
Duckworth, W.C. et al. (1998) Endocr. Rev. 19:608.
Akiyama, H. et al. (1990) Biochem. Biophys. Res. Commun. 170:1325.
Selkoe, D.J. (2001) Neuron 32:177.
Ghosh, S. et al. (2000) Am. J. Hum. Genet. 67:1174.
Farris, W. et al. (2003) Proc. Natl. Acad. Sci. USA 100:4162.

Long Name

Insulin Degrading Enzyme

Entrez Gene IDs

3416 (Human); 15925 (Mouse); 25700 (Rat)

Alternate Names

Abeta-degrading protease; EC 3.4.24; EC 3.4.24.56; FLJ35968; IDE; INSDEGM; Insulin protease; Insulinase; insulin-degrading enzyme; Insulysin

Related Research Areas

Citations for Recombinant Human Insulysin/IDE Protein, CF

R&D Systems personnel manually curate a database that contains references using R&D Systems products. The data collected includes not only links to publications in PubMed, but also provides information about sample types, species, and experimental conditions.

7 Citations: Showing 1 - 7
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Medium-Chain Length Fatty Acids Enhance Abeta Degradation by Affecting Insulin-Degrading Enzyme
Authors: J Mett, AA Lauer, D Janitschke, LV Griebsch, EL Theiss, HS Grimm, H Koivisto, H Tanila, T Hartmann, MOW Grimm
Cells, 2021-10-29;10(11):.
Species: Human
Sample Types: Recombinant Proteins
Applications: Bioassay
Discovery of O-glycans on atrial natriuretic peptide (ANP) that affect both its proteolytic degradation and potency at its cognate receptor
Authors: LH Hansen, TD Madsen, CK Goth, H Clausen, Y Chen, N Dzhoyashvi, SR Iyer, SJ Sangaralin, JC Burnett, JF Rehfeld, SY Vakhrushev, KT Schjoldage, JP Goetze
J. Biol. Chem., 2019-06-11;0(0):.
Species: Human
Sample Types: Whole Cells
Applications: Bioassay
The insulin-degrading enzyme is an allosteric modulator of the 20S proteasome and a potential competitor of the 19S
Authors: D Sbardella, GR Tundo, A Coletta, J Marcoux, EI Koufogeorg, C Ciaccio, AM Santoro, D Milardi, G Grasso, P Cozza, MP Bousquet-D, S Marini, M Coletta
Cell. Mol. Life Sci., 2018-03-28;0(0):.
Species: Rat
Sample Types: Proteasomes
Applications: Bioassay
Ex vivo (18)O-labeling mass spectrometry identifies a peripheral amyloid ? clearance pathway
Authors: E Portelius, N Mattsson, J Pannee, H Zetterberg, M Gisslén, H Vanderstic, E Gkanatsiou, GA Crespi, MW Parker, LA Miles, J Gobom, K Blennow
Mol Neurodegener, 2017-02-20;12(1):18.
Species: Human
Sample Types: CSF
Applications: Enzyme Assay
Characterization of insulin degrading enzyme and other amyloid-beta degrading proteases in human serum: a role in Alzheimer's disease?
Authors: Liu Z, Zhu H, Fang G, Walsh K, Mwamburi M, Wolozin B, Abdul-Hay S, Ikezu T, Leissring M, Qiu W
Oncogene, 2012-01-01;29(2):329-40.
Species: Human
Sample Types: Serum
Applications: Control
HIV-1 reduces Abeta-degrading enzymatic activities in primary human mononuclear phagocytes.
Authors: Lan X, Xu J, Kiyota T, Peng H, Zheng JC, Ikezu T
J. Immunol., 2011-05-06;186(12):6925-32.
Species: Virus
Sample Types: Protein
Applications: Enzyme Assay
Immunocapture-based fluorometric assay for the measurement of neprilysin-specific enzyme activity in brain tissue homogenates and cerebrospinal fluid.
Authors: Miners JS, Verbeek MM, Rikkert MO, Kehoe PG, Love S
J. Neurosci. Methods, 2007-08-25;167(2):229-36.
Species: Human
Sample Types: Peptide
Applications: Enzyme Assay