1 1

MMP-8 His Tag Protein, Cynomolgus

MMP-8 His Tag Protein, Cynomolgus

Catalog Number: UA010308 Brand: UA BIOSCIENCE
Share:
Price:
$840 USD
$840 USD
Size:

For shipping services or bulk orders, you may request a quotation.
Secure checkout with
View full details

Product Details

Product Specification


Species Cynomolgus
Synonyms MMP-8,CLG1,HNC,PMNL-CL
Accession XP_005579523.2
Amino Acid Sequence

Phe21-Ser467, with C-terminal 10*His
Expression System HEK293
Molecular Weight

65-75kDa
Purity >95% by SDS-PAGE
Endotoxin <0.1EU/μg
Conjugation Unconjugated
Tag His Tag
Physical Appearance Lyophilized powder
Storage Buffer

PBS, pH7.4
Reconstitution

Reconstitute at 0.1-1 mg/ml according to the size in ultrapure water after rapid centrifugation.
Stability & Storage

12 months from date of receipt, -20 to -70 °C as supplied; 6 months, -20 to -70 °C under sterile conditions after reconstitution; 1 week, 2 to 8 °C under sterile conditions after reconstitution; Please avoid repeated freeze-thaw cycles.
Reference

1、Hasty K A. et al. (1990) Human neutrophil collagenase. A distinct gene product with homology to other matrix metalloproteinases. J. Biol. Chem. 265: 11421-11424.

2、Blaeser J. et al. (1991) Mercurial activation of human polymorphonuclear leucocyte procollagenase. Eur. J. Biochem. 202: 1223-1230.

Background

A protease is an enzyme with a proteolytic activity that facilitates the digestion of its substrates. Membrane-type I matrix metalloproteinase (MT1-MMP), a member of the broader matrix metalloproteinases (MMP) family, is involved in the regulation of diverse cellular activities. Parasite-derived products have been shown to regulate host matrix metalloproteinases (MMPs), contributing to CE pathogenesis and progressive liver fibrosis in intermediate hosts. The current study aimed to investigate the potential role of MMP1, 7, 8, and 13 in E. granulosus induced liver fibrosis.MMP-8 may affect the metastatic behavior of breast cancer cells through protection against lymph node metastasis, underlining the importance of anti-target identification in drug development. MMP-8 in the tumor may have a protective effect against lymph node metastasis.

Protocol

Assay protocol

Principle: Measured by its ability to cleave a peptide substrate, Mca-PLGL-Dpa-AR-NH2.

Materials

  1. Assay Buffer: 50 mM Tris, 10 mM CaCl2, 150 mM NaCl, 0.05% (w/v) Brij 35, pH 7.5 (TCNB)

  2. MMP-8 His Tag Protein, Cynomolgus

  3. p-aminophenylmercuric acetate (APMA) (Aladdin, Catalog # P649286)

  4. Fluorogenic Peptide Substrate: MCA-Pro-Leu-Gly-Leu-DPA-Ala-Arg-NH2 (R&D, ES001)

  5. Calibration standard: MCA-Pro-Leu-OH (Shyuanye, T77046)

  6. 96 ELISA Removable Plate, Black, High binding (GENEVER, Catalog # GMO2-96H)

  7. Plate Reader (PerkinElmer, excitation 320 nm, and emission 405 nm)

Produce

  1. Activate MMP-8 at 100 μg/mL with 1 mM APMA in Assay Buffer. Incubate at 37 ℃ for 1 hour.

  2. Dilute activated MMP-8 (From the step 1) to 2, 1 and 0.5 μg/mL in Assay Buffer.

  3. Dilute Substrate to 20 μM in Assay Buffer.

  4. Load 50 μL of the dilute MMP-8 Protein (From the step 2) into a black well plate and start the reaction by adding 50 μL of 20 μM Substrate. Include a Substrate with Assay Buffer as Blank.

  5. Read at excitation and emission wavelengths of 320 nm and 405 nm, kinetic mode, 60s/cycle, 10 cycles.

  6. Calculate specific activity.

• Standard Curve

1. Dilute Calibration standard to 10 μM in Assay Buffer and prepare serial dilutions.

2. Add 100 μL of each serially diluted standard and blank (Assay buffer) into appropriate wells of a 96-well, the standard curve has a range of 500, 250, 125, 62.5, 31.25, 15.63, 7.81, 3.91, 1.95 pmol per well.

3. Read at excitation and emission wavelengths of 320 nm and 405 nm, respectively in endpoint mode.

4. Linear Regression of MCA-Pro-Leu-OH (pmol)(y) – RFU-Blank(x).

Specific Activity (pmol/min/µg) =

Adjusted Vmax (RFU/min) x Conversion Factor (pmol/RFU)

amount of enzyme (µg)

 

Picture

SDS-PAGE

1μg (R: reducing conditions, N: non-reducing conditions).