M9195

Matrix Metalloproteinase-1 human

recombinant, expressed in NSO cells, >95% (SDS-PAGE), buffered aqueous solution

• Synonyme(s) : Collagenase-1, Interstitial Collagenase, MMP-1
• Numéro de classification (Commission des enzymes): 3.4.24.7 (BRENDA, IUBMB)
• Numéro MDL: MFCD00130830
• Code UNSPSC : 12352202
• Nomenclature NACRES : NA.32

Propriétés

• Produit recombinant: expressed in NSO cells
• Niveau de qualité: 200
• Pureté: >95% (SDS-PAGE)
• Forme: buffered aqueous solution
• enzyme activity: ≥400 pmol/min-μg
• Poids mol.: apparent mol wt 52-55 kDa
• Numéro d'accès UniProt: P03956
• Conditions d'expédition: dry ice
• Température de stockage: −70°C
• Informations sur le gène: human ... MMP1(4312)

Description

Actions biochimiques/physiologiques

MMP-1 initiates the breakdown of interstitial collagens.

Forme physique

Supplied as a 0.2 μm filtered solution of 25 mM MES, 10 mM calcium chloride, 150 mM sodium chloride, 0.05% Brij^® L23, pH 5.5.

Remarque sur l'analyse

The biological activity is measured by its ability to cleave a fluorogenic peptide substrate.

Autres remarques

Activity Assay Protocol

Materials:

• Assay Buffer: 50 mM Tris, 10 mM CaCl2, 150 mM NaCl, 0.05% Brij-35, pH 7.5 (TCNB)
• Recombinant Human MMP-1 (rhMMP-1) (Cat. No. M9195)
• p-aminophenylmercuric acetate (APMA), (Cat. No. A9563), 100 mM stock in DMSO
• Substrate: MCA-Lys-Pro-Leu-Gly-Leu-DPA-Ala-Arg-NH2
• F16 Black Maxisorp Plate
• Fluorescent Plate Reader or equivalent

Assay:
1. Dilute rhMMP-1 to 50 μg/mL in Assay Buffer.
2. Activate 50 μg/mL rhMMP-1 by adding APMA to a final concentration of 1 mM.
3. Incubate at 37 °C for 2 hours.
4. Dilute activated rhMMP-1 to 1 ng/μL in Assay Buffer.
5. Dilute Substrate to 20 μM in Assay Buffer.
6. Load into a black well plate 50 μL of 1 ng/μL rhMMP-1 and start the reaction by adding 50 μL of 20 μM Substrate. Include a Substrate Blank containing 50 μL Assay Buffer, 50 μL Substrate, and no rhMMP-1.
7. Read at excitiation and emission wavelengths of 320 nm and 405 nm, respectively, in kinetic mode for 5 minutes.
8. 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.

Final Assay Conditions:
Per Well:
rhMMP-1: 0.050 μg
Substrate: 10 μM

Informations légales

Brij is a registered trademark of Croda International PLC

Informations sur la sécurité

• Code de la classe de stockage: 10 - Combustible liquids
• Classe de danger pour l'eau (WGK): WGK 2
• Point d'éclair (°F): Not applicable
• Point d'éclair (°C): Not applicable
• Équipement de protection individuelle: Eyeshields, Gloves, multi-purpose combination respirator cartridge (US)