Recombinant Human Neurolysin Protein, CF

Catalog #: 3814-ZN Datasheet / COA / SDS

Catalog #	Availability	Size / Price	Qty
3814-ZN-010

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Recombinant Human Neurolysin 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 a fluorogenic peptide substrate, (7-methoxycoumarin-4-yl)acetyl-Pro-Leu-Gly-Pro-D-Lys(2,4-dinitrophenyl)-OH or Mca-PLGPK(Dnp)-OH. The specific activity is >70 pmol/min/µg, as measured under the described conditions.

Source

E. coli-derived human Neurolysin protein
Ser38-Pro704, with an N-terminal Met and 6-His tag

Accession #

Q9BYT8

N-terminal Sequence
Analysis

Met

Predicted Molecular Mass

77 kDa

SDS-PAGE

73 kDa, reducing conditions

Product Datasheets

3814-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.

3814-ZN

Formulation	Supplied as a 0.2 μm filtered solution in Tris, NaCl and Glycerol.
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: 25 mM Tris, 150 mM NaCl, pH 7.5
Recombinant Human Neurolysin (rhNeurolysin) (Catalog # 3814-ZN)
Substrate: MCA-Pro-Leu-Gly-Pro-D-Lys(DNP)-OH (Bachem, Catalog # M-2270), 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 rhNeurolysin to 5 µg/mL in Assay Buffer.
Dilute Substrate to 50 µM in Assay Buffer.
Load into a black well plate 50 µL of 5 µg/mL rhNeurolysin, and start the reaction by adding 50 µL of 50 µM Substrate. Include a Substrate Blank containing 50 µL Assay Buffer and 50 µL of 50 µM Substrate.
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:

rhNeurolysin: 0.25 µg
Substrate: 25 µM

Reconstitution Calculator

Background: Neurolysin

Neurolysin, also known as Oligopeptidase M (mitochondrial peptidase), soluble angiotensin II-binding protein and endopeptidase EC 3.4.24.16 (1, 2), is a homologue of Thimet Ologopeptidase (THOP1), a zinc peptidase of the M3 family that also includes mitochondrial intermediate peptidase. Neurolysin expresses in two forms with the difference of an N-terminal transit peptide that targets the localization to mitochondria (1). The shorter form exists in cytosol. The recombinant human Neurolysin is expressed without the transit peptide and is isolated from the cytosol. Like THOP1, Neurolysin is capable of cleaving a number of vasoactive peptides such as bradykinin and neurotensin (3). All known substrates of THOP1 and Neurolysin contain 17 or fewer amino acids.

References

Serizawa, A. et al. (1995) J. Biol. Chem. 270:2092.
Barett, A.J. and J.M. Chen (2004) in Handbook of Proteolytic Enzymes (ed. Barrett, A.J. et al.) pp. 356, Elsevier Academic Press, San Diego.
Norman, M.U. et al. (2003) Am. J. Physiol. Heart Circ. Physiol. 284:H1978.

Entrez Gene IDs

57486 (Human); 75805 (Mouse); 117041 (Rat)

Alternate Names

AGTBP; AGTBPFLJ23002; angiotensin binding protein; Angiotensin-binding protein; DKFZp564F123; EC 3.4.24; EC 3.4.24.16; KIAA1226EP24.16; MEP; mitochondrial; MOP; neurolysin (metallopeptidase M3 family); Neurolysin; Neurotensin Endopeptidase; NLN; Oligopeptidase M