CHIT1/Chitotriosidase-1 Protein, Human (HEK293, His)

CHIT1/Chitotriosidase-1 protein, an enzyme degrading chitin, chitotriose, and chitobiose, potentially defends against nematodes and pathogens. Isoform 3 lacks enzymatic activity, highlighting the complexity of CHIT1 variants. The ability of CHIT1 to break down chitin suggests involvement in immune defense mechanisms. The existence of enzymatically inactive isoform 3 hints at diverse functions or regulatory roles within biological processes. CHIT1/Chitotriosidase-1 Protein, Human (HEK293, His) is the recombinant human-derived CHIT1/Chitotriosidase-1 protein, expressed by HEK293 , with C-His labeled tag.

The CHIT1/Chitotriosidase-1 protein serves as an enzyme capable of degrading chitin, chitotriose, and chitobiose, indicating its potential role in the defense against nematodes and other pathogens. However, it's noteworthy that isoform 3 of CHIT1 lacks enzymatic activity. The ability of CHIT1 to break down chitin-containing substances suggests its involvement in immune defense mechanisms, particularly against certain types of pathogens. The existence of isoform 3, which lacks enzymatic activity, underscores the complexity of CHIT1 protein variants and hints at potential diverse functions or regulatory roles within biological processes.

Measured by its ability to cleave 4-Methylumbelliferyl-beta-D-N,N’,N”-triacetylchitotriose (4-MU-TACT) that incubate at room temperature for 5 minutes. The specific activity is 2707 pmol/min/µg.

Materials
CHIT1/Chitotriosidase-1 Protein, Human (HEK293, His) (HY-P76254)
Assay buffer: 10 mM MES, pH 6.0
Reading Buffer: 100 mM Tris, pH 9.0
Substrate: 4-Methylumbelliferyl beta-D-N,N',N''-triacetylchitotriose
Standard: 4-Methylumbelliferone

Procedure
1. Dissolve the standard in reading buffer to prepare different concentration gradients with final concentrations of 5000, 2500, 1250, 625, 312.5, 156.25, 78, 39, 19, 9.75, and 0 pmol respectively. Read at excitation wavelength of 380 (10) nm and emission wavelength of 445 nm.
2. Dilute CHIT1 to 0.4 ng/μL with assay buffer.
3. Dilute the substrate to 200 μM with assay buffer.
4. Experimental group: Mix equal volumes of the test protein and substrate. Control group: Add equal volumes of assay buffer and substrate.
5. Incubate at room temperature for 5 minutes.
6. Add 90 μL of reading buffer to each well of the plate.
7. Add 10 μL of the reaction solution from the experimental group and the blank group to the above wells respectively.
8. Read in endpoint mode at excitation wavelength of 380 (10) nm and emission wavelength of 445 nm respectively.
9. Calculate the specific activity:

Human

HEK293

C-His

Q13231 (A22-N466)

1118  [NCBI]

Full Length of Mature Protein

AKLVCYFTNWAQYRQGEARFLPKDLDPSLCTHLIYAFAGMTNHQLSTTEWNDETLYQEFNGLKKMNPKLKTLLAIGGWNFGTQKFTDMVATANNRQTFVNSAIRFLRKYSFDGLDLDWEYPGSQGSPAVDKERFTTLVQDLANAFQQEAQTSGKERLLLSAAVPAGQTYVDAGYEVDKIAQNLDFVNLMAYDFHGSWEKVTGHNSPLYKRQEESGAAASLNVDAAVQQWLQKGTPASKLILGMPTYGRSFTLASSSDTRVGAPATGSGTPGPFTKEGGMLAYYEVCSWKGATKQRIQDQKVPYIFRDNQWVGFDDVESFKTKVSYLKQKGLGGAMVWALDLDDFAGFSCNQGRYPLIQTLRQELSLPYLPSGTPELEVPKPGQPSEPEHGPSPGQDTFCQGKADGLYPNPRERSSFYSCAAGRLFQQSCPTGLVFSNSCKCCTWN

Approximately 48 kDa.

≥ 90%, as determined by reducing SDS-PAGE.

Solution.

Supplied as a 0.22 μm filtered solution of 12.5 mM Tris, 75 mM NaCl, 50% glycerol, pH 7.5.
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.

N/A.

Stored at -80°C for 1 year from date of receipt. It is stable at -20°C for 3 months after opening. It is recommended to freeze aliquots at -80°C for extended storage. Avoid repeated freeze-thaw cycles.

Shipping with dry ice.

• [1]. /biology-dictionary/hek293-cell.html  [Content Brief]