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Recombinant Human Neprilysin (CHO-expressed) Protein, CF

Catalog #: 1182-ZNC
6 Citations Datasheet
Catalog # Availability Size / Price Qty
1182-ZNC-010
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Citations (6)
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Recombinant Human Neprilysin (CHO-expressed) 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 >1,500 pmol/min/µg, as measured under the described conditions.
Source
Chinese Hamster Ovary cell line, CHO-derived human Neprilysin/CD10 protein
Tyr52-Trp750, with an N-terminal 6-His tag
Accession #
P08473
N-terminal Sequence
Analysis
His
Predicted Molecular Mass
81 kDa
SDS-PAGE
102 kDa, reducing conditions

Product Datasheets

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1182-ZNC

Product Datasheet

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.

1182-ZNC

Formulation Supplied as a 0.2 μm filtered solution in Tris, NaCl and ZnCl2.
Shipping The product is shipped with polar packs. 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, 0.05% Brij-35, pH 9.0
  • Recombinant Human Neprilysin (rhNeprilysin) (Catalog # 1182-ZNC)
  • Substrate: MCA-Arg-Pro-Pro-Gly-Phe-Ser-Ala-Phe-Lys(DNP)-OH (Catalog # ES005), Prepare 2 mM stock in DMSO
  • F16 Black Maxisorp Plate (Nunc, Catalog # 475515)
  • Fluorescent Plate Reader (Model: SpectraMax Gemini EM by Molecular Devices) or equivalent
  1. Dilute rhNeprilysin to 0.1 µg/mL in Assay Buffer.
  2. Dilute fluorogenic peptide Substrate to 20 µM in Assay Buffer.
  3. Load in a black well plate 50 µL of 0.1 µg/mL of rhNeprilysin (5 ng/well), and start the reaction by adding 50 µL of 20 µM Substrate. As a Substrate Blank combine 50 µL of Substrate and 50 µL of Assay Buffer.
  4. Read at excitation and emission wavelengths of 320 nm and 405 nm (top read), respectively in kinetic mode for 5 minutes.
  5. Calculate specific activity:
     Specific Activity (pmol/min/µg) =
 Adjusted Vmax* (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:
  • rhNeprilysin: 0.005 µg
  • Substrate: 10 µM

Background: Neprilysin/CD10

Neprilysin/CD10, also known as NEP and neutral endopeptidase 24.11, is a zinc metallopeptidase expressed at the cell surface of a variety of cells. The enzyme functions both as an endopeptidase with a thermolysin-like specificity and as a dipeptidylcarboxypeptidase. NEP has been shown to be involved in the degradation of enkephalins in the mammalian brain and the inactivation of circulating atrial natriuretic peptide (1, 2). NEP has also been identified as the common acute lymphocytic leukemia antigen (CALLA), and is expressed on the surface of lymphocytes in some disease states (3, 4). These and other observations have resulted in considerable interest in NEP as a target for analgesics and antihypertensive drugs. NEP is also a major degrading enzyme of amyloid beta  peptide (A beta ) in the brain, indicating that down-regulation of NEP activity, which could be caused by aging, can contribute to the development of Alzheimer’s disease by promoting A beta  accumulation (5).

References
  1. Malfroy, B. et al. (1978) Nature 276:523.
  2. Kenny, A.J. and Stephenson, S.L. (1988) FEBS Lett. 232:1.
  3. LeTarte, M. et al. (1988) J. Exp. Med. 168:1247.
  4. Shipp, M.A. et al. (1988) Proc. Natl. Acad. Sci. USA 85:4819.
  5. Itwata, N. et al. (2001) Science 292:1550.
Entrez Gene IDs
4311 (Human); 17380 (Mouse); 24590 (Rat); 101925772 (Cynomolgus Monkey)
Alternate Names
Atriopeptidase; CALLA; CALLAmembrane metallo-endopeptidase (neutral endopeptidase, enkephalinase); CD10 antigen; CD10; CD10); CD10membrane metallo-endopeptidase variant 1; Common acute lymphocytic leukemia antigen; DKFZp686O16152; EC 3.4.24; EC 3.4.24.11; Enkephalinase; EPN; Leu-19; membrane metallo-endopeptidase variant 2; membrane metallo-endopeptidase; MGC126681; MGC126707; MME; NEPmembrane metallo-endopeptidase (neutral endopeptidase, enkephalinase, CALLA; Neprilysin; Neutral Endopeptidase 24.11; Neutral endopeptidase; NKH1; SFE; Skin fibroblast elastase

Citations for Recombinant Human Neprilysin (CHO-expressed) 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.

6 Citations: Showing 1 - 6
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  1. Comparative studies for amyloid beta degradation: "Neprilysin vs insulysin", "monomeric vs aggregate", and "whole Abeta40 vs its peptide fragments"
    Authors: D Kato, Y Takahashi, H Iwata, Y Hatakawa, SH Lee, T Oe
    Biochemistry and Biophysics Reports, 2022-05-09;30(0):101268.
    Species: Human
    Sample Types: Peptides
    Applications: Bioassay
  2. 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
  3. Discovery of a long-acting glucagon-like peptide-1 analog with enhanced aggregation propensity
    Authors: M Takeuchi, M Okamoto, R Okamoto, H Kinoshita, Y Yamaguchi, N Watanabe
    Peptides, 2018-01-31;102(0):8-15.
    Species: Mouse
    Sample Types: Serum
    Applications: Bioassay
  4. Degradation of tropoelastin and skin elastin by neprilysin
    Authors: AC Mora Huert, CEH Schmelzer, C Luise, W Sippl, M Pietzsch, W Hoehenwart, A Heinz
    Biochimie, 2017-11-29;146(0):73-78.
    Species: Human
    Sample Types: Whole Cells
    Applications: Cell Culture
  5. Lentivirus mediated delivery of neurosin promotes clearance of wild-type alpha-synuclein and reduces the pathology in an alpha-synuclein model of LBD.
    Authors: Spencer B, Michael S, Shen J, Kosberg K, Rockenstein E, Patrick C, Adame A, Masliah E
    Mol Ther, 2012-04-17;21(1):31-41.
    Applications: Bioassay
  6. 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

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