Kallikrein-4 Protein, Human (H197Q, HEK293, His)

Kallikrein-4 Protein is crucial in enamel formation, particularly during tooth maturation, playing a significant role in clearing enamel proteins and contributing to the structural patterning of the crystalline matrix. Its involvement underscores its importance in intricate tooth development processes, emphasizing its role in proper dental structure and function. Kallikrein-4 Protein, Human (H197Q, HEK293, His) is the recombinant human-derived Kallikrein-4 protein, expressed by HEK293 , with C-6*His labeled tag and H197Q mutation.

Kallikrein-4 Protein plays a significant role in enamel formation, particularly during the maturation stage of tooth development. It is essential for the clearance of enamel proteins and contributes to the normal structural patterning of the crystalline matrix. This underscores the protein's crucial involvement in the intricate processes underlying tooth development and enamel maturation, highlighting its importance for proper dental structure and function.

Measured by its ability to cleave 100μM fluorogenic peptide substrate Boc-VPR-AMC (HY-137784) that incubate at 37℃ in kinetic mode for 5 minutes. The specific activity is 412.94 pmol/min/µg. (Activation description: The proenzyme needs to be activated by Thermolysin for an activated form.)

Materials
Activation buffer: 50 mM Tris, 150 mM NaCl, 10 mM CaCl₂, pH 7.5
Assay buffer: 50 mM Tris, pH 9.0
Kallikrein-4 Protein, Human (H197Q, HEK293, His) (HY-P70947)
Substrate: Boc-Val-Pro-Arg-AMC hydrochloride (HY-137784), dissolved in DMSO to 10 mM

Procedure
1. Prepare standards with assay buffer to final concentrations of 0, 9.76, 19, 39, 78, 156, 312.5, 625, 1250, 2500, 5000, 10000 pmoL respectively, and read at excitation and emission wavelengths of 380 nm and 460 nm.
2. Dilute Human Kallikrein-4 to 200 μg/mL with activation buffer.
3. Dilute thermolysin to 2 μg/mL with activation buffer.
4. Mix equal volumes of the above test protein and thermolysin, and incubate at 37℃ for 2 h.
5. Terminate with 1,10-Phenanthroline at a final concentration of 10 mM.
6. Dilute the activated test protein to 2 μg/mL with assay buffer.
7. Dilute the substrate to 200 μM with assay buffer.
8. Add 50 μL of activated test protein to the wells of a black well plate, and add 50 μL of 200 μM substrate to each well to start the reaction. For blank wells, add 50 μL assay buffer and 50 μL of 200 μM substrate.
9. Read in kinetic mode for 5 minutes at excitation and emission wavelengths of 380 nm and 460 nm respectively.
10. Calculate the specific activity:

Human

HEK293

C-6*His

Q9Y5K2 (S27-S254, H197Q)

9622  [NCBI]

Full Length of Mature Protein

SCSQIINGEDCSPHSQPWQAALVMENELFCSGVLVHPQWVLSAAHCFQNSYTIGLGLHSLEADQEPGSQMVEASLSVRHPEYNRPLLANDLMLIKLDESVSESDTIRSISIASQCPTAGNSCLVSGWGLLANGRMPTVLQCVNVSVVSEEVCSKLYDPLYHPSMFCAGGGQDQKDSCNGDSGGPLICNGYLQGLVSFGKAPCGQVGVPGVYTNLCKFTEWIEKTVQAS

25-32 kDa

• 
  Greater than 95% as determined by reducing SDS-PAGE.

Lyophilized powder.

Lyophilized from a 0.22 μm filtered solution of 20 mM Tris-HCl, 150 mM NaCl, pH 8.0.
Note: For SPR assay, please replace the buffer. Primary amine components (e.g., Tris, imidazole) can affect protein-coupled chips.

<1 EU/μg, determined by LAL method.

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).

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.

Room temperature in continental US;may vary elsewhere.