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Cathepsin B Protein, Mouse (HEK293, His)

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Based on 2 publication(s) in Google Scholar

Cathepsin B Protein, Mouse (HEK293, His) is an approximately 32-50 kDa Cathepsin B protein with a His-flag. Cathepsin B is an enzymatic protein belonging to the peptidase C1 family.

For research use only. We do not sell to patients.
  • Species: Mouse
  • Source: HEK293
  • Storage:
    Stored at -20°C for 2 years from date of receipt. After reconstitution, it is stable at 4°C for 1 week or -20°C for longer (with carrier protein). It is recommended to freeze aliquots at -20°C or -80°C for extended storage.
  • Experimental Validation Results for Cathepsin B Protein, Mouse (HEK293, His)
  • Cathepsin B Protein, Mouse (HEK293, His) Product Image
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[email protected] [email protected]
Find Your Local Distributor Your local distributor: Hycultec GmbHHycultec
  • Biological Activity
  • Technical Parameters
  • Product Properties
  • Documentation
  • References
  • Help & FAQs
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Use of MCE Products

Biological Activity

Description

Cathepsin B Protein, Mouse (HEK293, His) is an approximately 32-50 kDa Cathepsin B protein with a His-flag. Cathepsin B is an enzymatic protein belonging to the peptidase C1 family[1].

Background

Cathepsin B (CTSB) is also known as APP secretase (APPS) and CPSB, is an enzymatic protein belonging to the peptidase C1 family[1].
Cathepsin B/CTSB is synthesized as a preproenzyme and is activated in prelysosomal acidic vesicles (late endosomes) prior to its delivery to lysosomes. After removal of the signal peptide, the inactive proenzyme undergoes modifications including removal of the pro region to result in the active enzyme[1].
Several lysosomal proteinases including the cathepsin B have been implicated in malignant progression of tumors. Many researchers have demonstrated correlations between increased activity of cathepsin B and increased metastatic capability of animal tumors or malignancy of human tumors[2].

Verified Bioactivity

Measured by its ability to cleave 10 μM fluorogenic peptide 10 μM substrate Z-LR-AMC (HY-142021). The specific activity is >2000 pmol/min/µg, as measured under the described conditions. (Activation description: The proenzyme needs to be activated in acid reducing buffer for an activated form.)

MCE Validation Data

  • Purity - SDS-PAGE

    Purity - SDS-PAGE

    ≥ 95%, as determined by reducing SDS-PAGE.

Assay Procedure

Materials
Activation Buffer: 25 mM MES, 5 mM DTT, pH 5.0.
Assay Buffer: 25 mM MES, pH 5.0.
Test protein: Cathepsin B Protein, Mouse (HEK293, His) (HY-P7747)
Standard: 7-Amino, 4-Methyl Coumarin (AMC, HY-D0027)
Substrate: Z-Leu-Arg-AMC (Z-LR-AMC, HY-142021)

Procedure
1. Standard curve: Dilute 7-Amino, 4-Methyl Coumarin (AMC) (standard) to 0, 0.78125, 1.5625, 3.125, 6.25, 12.5, 25, 50, 100 μmol/L with assay buffer, and add 100 μL into the black enzyme-labeled wells. The excitation and emission wavelengths were 380 nm and 460 nm, respectively, and the standard curve was made with the measured RFU as the horizontal coordinate and the amount of standard substance as the vertical coordinate to obtain the standard curve formula.
2. Dilute the measured proteins to 10μg/mL with activation buffer.
3. Incubate for 15 minutes at room temperature.
4. Dilute Mouse Cathepsin B to 0.2 μg/mL in Assay Buffer.
5. Dilute substrate to 20 μM in assay buffer.
6. Add 50 μL of each concentration of the measured protein to the assay wells and add 50 μL of 20 μM substrate to start the reaction. Blank wells were spiked with 50 μL of assay buffer and 50 μL of 20μM protein-free substrate.
7. The excitation and emission wavelengths were 380 nm and 460 nm, respectively, and read in kinetic mode for 5 minutes.
8. Calculate the specific activity:

     Specific Activity (pmol/min/μg) =

Adjusted Vmax* (RFU/min) x Conversion Factor ** (pmol/RFU)
amount of enzyme (μg)

*Adjusted for Control
**Derived using calibration standard

Per Well:
Mouse Cathepsin B: 0.01 μg
Substrate: 10 μM

Technical Parameters

  • Species Mouse
  • Source HEK293
  • Tag C-6*His
  • Accession

    P10605 (H18-F339)

  • Gene ID
    13030  [NCBI]
  • Molecular Construction
    • N-term
    • Cathepsin B (H18-F339)
      Accession # P10605
    • 6*His
    • C-term
  • Protein Length

    Full Length of Mature Protein

  • Synonyms

    CTSB; Keratolytic Winter Erythema (Oudtshoorn Skin Disease); Cathepsin B; Epididymis Secretory Sperm Binding Protein; APP Secretase; Amyloid Precursor Protein Secretase; Cathepsin B1; Cysteine Protease; APPS; RECEUP; CPSB; KWE

  • AA Sequence

    HDKPSFHPLSDDLINYINKQNTTWQAGRNFYNVDISYLKKLCGTVLGGPKLPGRVAFGEDIDLPETFDAREQWSNCPTIGQIRDQGSCGSCWAFGAVEAISDRTCIHTNGRVNVEVSAEDLLTCCGIQCGDGCNGGYPSGAWSFWTKKGLVSGGVYNSHVGCLPYTIPPCEHHVNGSRPPCTGEGDTPRCNKSCEAGYSPSYKEDKHFGYTSYSVSNSVKEIMAEIYKNGPVEGAFTVFSDFLTYKSGVYKHEAGDMMGGHAIRILGWGVENGVPYWLAANSWNLDWGDNGFFKILRGENHCGIESEIVAGIPRTDQYWGRF

  • Molecular Weight

    Approximately 43-47 kDa & 32-35 kDa, based on SDS-PAGE under reducing conditions, due to the glycosylation.

  • Glycosylation

    Yes

  • Purity

    ≥ 95%, as determined by reducing SDS-PAGE.

Product Properties

Appearance

Lyophilized powder

Formulation

Lyophilized from a 0.22 μm filtered solution of PBS, pH 7.4.

Endotoxin Level

<1 EU/μg, determined by LAL method.

Reconstitution

It is not recommended to reconstitute to a concentration less than 100 μg/mL in ddH2O. For long term storage it is recommended to add a carrier protein (0.1% BSA, 5% HSA, 10% FBS or 5% Trehalose).

Storage & Stability

Stored at -20°C for 2 years from date of receipt. After reconstitution, it is stable at 4°C for 1 week or -20°C for longer (with carrier protein). It is recommended to freeze aliquots at -20°C or -80°C for extended storage.

Shipping

Room temperature in continental US; may vary elsewhere.

References

[1]. Izabela Podgorski, et al. Cathepsin B and its role(s) in cancer progression. Biochem Soc Symp. 2003;(70):263-76. [Content Brief]

[2]. W Morikawa, et al. Angiostatin generation by cathepsin D secreted by human prostate carcinoma cells. J Biol Chem. 2000 Dec 8;275(49):38912-20. [Content Brief]

Calculators

Reconstitution Calculator

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Dilution Calculator

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The Specific Activity Calculator Equation
  • Specific Activity (Unit/mg)
  • Biological Activity (ED50)

Specific Activity (Unit/mg) = 106 ÷ Biological Activity (ED50)

Specific Activity (Unit/mg) Specific Activity (Unit/mg)
Unit/mg
= 106 ÷
Biological Activity (ED50) Biological Activity (ED50)
106 ÷
ng/mL
MOQ
Minimum order quantity
100 mg

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Measured by its ability to inhibit anti-CD3-induced proliferation of stimulated CTLL-2 mouse cytotoxic T cells. The ED50 this effect is 0.8023 μg/ml, corresponding to a specific activity is 1246.42 units/mg.

Assay Types
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    MedChemExpress Validation
     ≥ 95%, as determined by reducing SDS-PAGE.

  • Cathepsin B Protein, Mouse (HEK293, His) Product Image
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  • Experimental Validation Results for Cathepsin B Protein, Mouse (HEK293, His)
  • Cathepsin B Protein, Mouse (HEK293, His) Product Image