Recombinant Human ADAM9 Protein, CF

• Reactivity: Hu
• Applications: Enzyme Activity
• Format: Carrier-Free

Recombinant Human ADAM9 Protein, CF Summary

• Details of Functionality: Measured by its ability to cleave a fluorogenic peptide substrate Mca-PLAQAV-Dpa-RSSSR-NH₂ (Catalog # ES003). The specific activity is >1.0 pmol/min/µg, as measured under the described conditions.
• Source: Mouse myeloma cell line, NS0-derived human ADAM9 protein
  Ala206-Asp697, with a C-terminal 10-His tag
• Accession #: Q13443
• N-terminal Sequence: Ala206
• Structure / Form: Mature form. Recombinant Human ADAM9 is prone to proteolytic cleavage at C-terminus. The poly-His tag may not be present in the preparation.
• Protein/Peptide Type: Recombinant Enzymes
• Gene: ADAM9
• Purity: >90%, by SDS-PAGE under reducing conditions and visualized by silver stain.
• Endotoxin Note: <1.0 EU per 1 μg of the protein by the LAL method.

Applications/Dilutions

• Theoretical MW: 55 kDa.
  Disclaimer note: The observed molecular weight of the protein may vary from the listed predicted molecular weight due to post translational modifications, post translation cleavages, relative charges, and other experimental factors.
• SDS-PAGE: 64 kDa, reducing conditions

Packaging, Storage & Formulations

• 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.
• Buffer: Supplied as a 0.2 μm filtered solution in MES and NaCl.
• Purity: >90%, by SDS-PAGE under reducing conditions and visualized by silver stain.
• Assay Procedure:
  • Assay Buffer: 25 mM Tris, 2.5 µM ZnCl₂, 0.005% (w/v) Brij-35, pH 9.0
  • Recombinant Human ADAM-9 (rhADAM9) (Catalog # 939-AD)
  • Fluorogenic Peptide Substrate III: MCA-Pro-Leu-Ala-Gln-Ala-Val-DPA-Arg-Ser-Ser-Ser-Arg-NH₂ (Catalog # ES003)
  • F16 Black Maxisorp Plate (Nunc, Catalog # 475515)
  • Fluorescent Plate Reader (Model: SpectraMax Gemini EM by Molecular Devices) or equivalent
  1. Dilute rhADAM9 to 40 µg/mL in Assay Buffer.
  2. Dilute substrate to 20 µM in Assay Buffer.
  3. Combine equal volumes of 40 µg/mL rhADAM9 and 20 µM substrate. Include a control containing Assay Buffer and 20 µM substrate without any rhADAM9.
  4. Incubate reactions at 37 °C for 30 minutes.
  5. Load 100 μL of reactions and control per well in a black plate.
  6. Read with excitation and emission wavelengths of 320 nm and 405 nm (top read), respectively, in endpoint mode.
  7. Calculate specific activity:

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

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

  Per Well:
  • rhADAM9: 2.0 µg
  • Substrate: 10 µM

Notes

This product is produced by and ships from R&D Systems, Inc., a Bio-Techne brand.

Alternate Names for Recombinant Human ADAM9 Protein, CF

• a disintegrin and metalloproteinase domain 9 (meltrin gamma)
• ADAM 9
• ADAM metallopeptidase domain 9 (meltrin gamma)
• ADAM metallopeptidase domain 9
• ADAM9
• Cellular disintegrin-related protein
• cone rod dystrophy 9
• CORD9
• disintegrin and metalloproteinase domain-containing protein 9
• EC 3.4.24
• EC 3.4.24.-
• MCMP
• MCMPMDC9KIAA0021Mltng
• MDC9
• Meltrin gamma
• Meltrin-gamma
• Metalloprotease/disintegrin/cysteine-rich protein 9
• MLTNG
• Myeloma cell metalloproteinase

Background

ADAM9, also known as MDC9 or meltrin gamma, is a member of the ADAM family that contains a disintegrin and metalloprotease‑like domain (1). Like other membrane-anchored ADAMs, ADAM9 consists of a pro domain with a cysteine switch and furin cleavage sequence, a catalytic domain with the zinc‑binding site and Met-turn expected for reprolysins, a disintegrin-like domain, a cysteine-rich domain, an EGF-like domain, a transmembrane domain, and the cytoplasmic domain. ADAM9 is able to cleave peptides corresponding to cleavage sites of tumor necrosis factor-alpha (TNF-alpha ), the p75 TNF receptor, the beta -amyloid protein precursor, and the c‑kit ligand-1, implying that it may participate in shedding of these membrane proteins (2). In fact, ADAM9 has been shown to shed membrane‑anchored heparin-binding EGF-like growth factor (3). In addition, it also cleaves oxidized insulin beta -chain and fibronectin (2,4). Besides its catalytic activity, ADAM9 functions as an adhesion molecule through binding of its disintegrin domain to integrins such as alpha _v beta ₅ and alpha ₆ beta ₁ (5, 6). The cytoplasmic domain of ADAM9 interacts with Src homology 3
(SH3)‑containing proteins and protein kinase C, and may mediate different signaling pathways (3, 7). ADAM9 is widely expressed in tissues (8).

1. Moss, et al. (2001) DDT 6:417.
2. Roghan, et al. (1999) J. Biol. Chem. 274:3531.
3. Izumi, et al. (1998) EMBO J. 17:7260.
4. Schwettmann and Tschesche (2001) Prot. Expre. & Purif. 21:65.
5. Nath, et al. (2000) J. Cell Sci. 113:2319.
6. Zhou, et al. (2001) Biochem. Biophys. Res. Comm. 280:574.
7. Howard, et al. (1999) J. Biol. Chem. 274:31693.
8. Weskamp, et al. (1996) J. Cell. Biol. 132:717.

Publications for ADAM9 (939-AD)(7)

Publications using 939-AD

• N Giebeler, A Schönefu beta, J Landsberg, T Tüting, C Mauch, P Zigrino Deletion of ADAM-9 in HGF/CDK4 mice impairs melanoma development and metastasis Oncogene, 2017;0(0):. 2017 [PMID: 28553955] (EnzAct, N/A)
• Schlomann U, Koller G, Conrad C, Ferdous T, Golfi P, Garcia A, Hofling S, Parsons M, Costa P, Soper R, Bossard M, Hagemann T, Roshani R, Sewald N, Ketchem R, Moss M, Rasmussen F, Miller M, Lauffenburger D, Tuveson D, Nimsky C, Bartsch J ADAM8 as a drug target in pancreatic cancer. Nat Commun, 2015;6(0):6175. 2015 [PMID: 25629724] (Primate - Chlorocebus aethiops (African Green Monkey))
• Roychaudhuri R, Hergrueter A, Polverino F, Laucho-Contreras M, Gupta K, Borregaard N, Owen C ADAM9 is a novel product of polymorphonuclear neutrophils: regulation of expression and contributions to extracellular matrix protein degradation during acute lung injury. J Immunol, 2014;193(5):2469-82. 2014 [PMID: 25063875] (EnzAct, Human)
• Moss ML, Bomar M, Liu Q, Sage H, Dempsey P, Lenhart PM, Gillispie PA, Stoeck A, Wildeboer D, Bartsch JW, Palmisano R, Zhou P The ADAM10 prodomain is a specific inhibitor of ADAM10 proteolytic activity and inhibits cellular shedding events. J. Biol. Chem., 2007;282(49):35712-21. 2007 [PMID: 17895248] (Bioassay, Human)
• Zhou BB, Peyton M, He B, Liu C, Girard L, Caudler E, Lo Y, Baribaud F, Mikami I, Reguart N, Yang G, Li Y, Yao W, Vaddi K, Gazdar AF, Friedman SM, Jablons DM, Newton RC, Fridman JS, Minna JD, Scherle PA Targeting ADAM-mediated ligand cleavage to inhibit HER3 and EGFR pathways in non-small cell lung cancer. Cancer Cell, 2006;10(1):39-50. 2006 [PMID: 16843264] (Enzyme Assay, Human)
• Karadag A, Zhou M, Croucher PI ADAM-9 (MDC-9/meltrin-gamma), a member of the a disintegrin and metalloproteinase family, regulates myeloma-cell-induced interleukin-6 production in osteoblasts by direct interaction with the alpha(v)beta5 integrin. Blood, 2006;107(8):3271-8. 2006 [PMID: 16373656] (Bioassay, Human)
• Hinkle CL, Sunnarborg SW, Loiselle D, Parker CE, Stevenson M, Russell WE, Lee DC Selective roles for tumor necrosis factor alpha-converting enzyme/ADAM17 in the shedding of the epidermal growth factor receptor ligand family: the juxtamembrane stalk determines cleavage efficiency. J. Biol. Chem., 2004;279(23):24179-88. 2004 [PMID: 15066986] (EnzAct, Mouse)

Bioinformatics

• Gene Symbol: ADAM9
• Entrez:
  • 8754(Human)
  • 11502(Mouse)
• Uniprot:
  • Q13443()