Recombinant Human Neprilysin (CHO-expressed) Protein, CF

Catalog #: 1182-ZNC
6 Citations Datasheet / COA / SDS

Catalog #	Availability	Size / Price	Qty
1182-ZNC-010

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Citations (6)

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

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

1182-ZNC

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.

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

Dilute rhNeprilysin to 0.1 µg/mL in Assay Buffer.
Dilute fluorogenic peptide Substrate to 20 µM in Assay Buffer.
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.
Read at excitation and emission wavelengths of 320 nm and 405 nm (top read), 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:

rhNeprilysin: 0.005 µg
Substrate: 10 µM

Reconstitution Calculator

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

Malfroy, B. et al. (1978) Nature 276:523.
Kenny, A.J. and Stephenson, S.L. (1988) FEBS Lett. 232:1.
LeTarte, M. et al. (1988) J. Exp. Med. 168:1247.
Shipp, M.A. et al. (1988) Proc. Natl. Acad. Sci. USA 85:4819.
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

Related Research Areas

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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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
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
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
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
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
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