Recombinant Human QPCT Protein, CF

• Reactivity: Hu
• Applications: Enzyme Activity
• Format: Carrier-Free

Recombinant Human QPCT Protein, CF Summary

• Details of Functionality: Measured by its ability to convert Glutaminyl-AMC to pyroglutamyl-AMC. The specific activity is >550 pmol/min/μg, as measured under the described conditions.
• Source: Spodoptera frugiperda, Sf 21 (baculovirus)-derived human Glutaminyl-peptide Cyclotransferase/QPCT protein
  Ala33-Leu361, with an N-terminal 6-His tag
• Accession #: Q16769
• N-terminal Sequence: His
• Protein/Peptide Type: Recombinant Enzymes
• Gene: QPCT
• Purity: >95%, by SDS-PAGE visualized with Silver Staining and quantitative densitometry by Coomassie® Blue Staining.
• Endotoxin Note: <1.0 EU per 1 μg of the protein by the LAL method.

Applications/Dilutions

• Dilutions:
  • Enzyme Activity
• Theoretical MW: 38 kDa.
  Disclaimer note: The observed molecular weight of the protein may vary from the listed predicted molecular weight due to post translational modifications, post translation cleavages, relative charges, and other experimental factors.
• SDS-PAGE: 36-40 kDa, reducing conditions

Packaging, Storage & Formulations

• Storage: Use a manual defrost freezer and avoid repeated freeze-thaw cycles.
  • 6 months from date of receipt, -20 to -70 °C as supplied.
  • 3 months, -20 to -70 °C under sterile conditions after opening.
• Buffer: Supplied as a 0.2 μm filtered solution in Tris, NaCl and Glycerol.
• Purity: >95%, by SDS-PAGE visualized with Silver Staining and quantitative densitometry by Coomassie® Blue Staining.
• Assay Procedure:
  • Assay Buffer: 25 mM HEPES, pH 7.0
  • rhPGPEP-1 Buffer: 0.1 M Tris, 5 mM DTT, pH 9.0
  • Recombinant Human Glutaminyl‑peptide Cyclotransferase/QPCT (rhQPCT) (Catalog # 6368-ZN)
  • Substrate: Q-AMC (Bachem, Catalog # I-1175)
  • Recombinant Human PGPEP-1 (rhPGPEP-1) (Catalog # 6278-CY)
  • F16 Black Maxisorp Plate (Nunc, Catalog # 475515)
  • Fluorescent Plate Reader (Model: SpectraMax Gemini EM by Molecular Devices) or equivalent
  1. Dilute rhQPCT to 0.2 μg/mL in Assay Buffer.
  2. Dilute Substrate to 200 μM in Assay Buffer.
  3. Combine 150 μL of 0.2 ug/mL rhQPCT and 150 μL of 200 μM Substrate. Include a Substrate Blank containing 150 μL Assay Buffer and 150 μL of 200 μM Substrate.
  4. Incubate reaction at room temperature for 20 minutes.
  5. Stop reaction by boiling at 100 °C for 5 minutes and cool on ice for 3 minutes.
  6. Dilute rhPGPEP-1 to 1 μg/mL in rhPGPEP-1 Buffer.
  7. Add 300 μL of 1 μg/mL rhPGPEP-1 to each vial.
  8. Incubate reaction at room temperature for 10 minutes.
  9. Load 100 μL in quadruplicate from each vial into a 96 well plate.
  10. Read plate at excitation and emission wavelengths of 380 nm and 460 nm, respectively, in endpoint mode.
  11. Calculate specific activity:

  Specific Activity (pmol/min/µg) = (Adjusted Fluorescence* (RFU) x Conversion Factor** (pmol/RFU)) / (Incubation time (min) x amount of enzyme (µg))

  *Adjusted for Substrate Blank
  **Derived using calibration standard 7-Amino, 4-Methyl Coumarin (Sigma, Catalog # A-9891).

  Per Well:
  • rhQPCT: 0.005 μg
  • Substrate: 50 μM

Notes

This product is produced by and ships from R&D Systems, Inc., a Bio-Techne brand.

Alternate Names for Recombinant Human QPCT Protein, CF

• EC 2.3.2.5
• GCT
• Glutaminyl cyclase
• glutaminyl-peptide cyclotransferase
• Glutaminyl-tRNA cyclotransferase
• Glutamyl cyclase
• QCEC
• QPCT
• sQC

Background

Glutaminyl-peptide Cyclotransferase, also known as Glutaminyl Cyclase, catalyzes the conversion of N‑terminal L-glutaminyl residues of peptides to pyroglutamyl groups (1). The enzyme is present in the pituitary and adrenal glands, where it is important for the generation of the N‑terminal pyroglutamyl groups of peptide hormones such as neurotensin and thyrotropin-releasing hormone. Glutaminyl Cyclase also catalyzes the conversion of N‑terminal L‑glutamyl residues to pyroglutamyl residues (2). This activity may contribute to the formation of several amyloid-related plaque forming peptides, contributing to Alzheimer’s disease pathology. Glutaminyl Cyclase is also considered to be a diagnostic marker of thyroid tumors (3).

1. Busby, W.H. Jr. et al. (1987) J. Biol. Chem. 262:8532.
2. Schilling, S. et al. (2004) FEBS Lett. 563:191.
3. Griffith, O.L. et al. (2006) J. Clin. Oncol. 24:5043.

Publications for Glutaminyl-peptide Cyclotransferase/QPCT (6368-ZN)(2)

Publications using 6368-ZN

• N Van Manh, VH Hoang, VTH Ngo, S Kang, JJ Jeong, HJ Ha, H Kim, YH Kim, J Ann, J Lee Discovery of potent indazole-based human glutaminyl cyclase (QC) inhibitors as Anti-Alzheimer&#039;s disease agents European Journal of Medicinal Chemistry, 2022-10-12;244(0):114837. 2022-10-12 [PMID: 36265279] (Bioassay, Human)
• N Van Manh, VH Hoang, VTH Ngo, J Ann, TH Jang, JH Ha, JY Song, HJ Ha, H Kim, YH Kim, J Lee, J Lee Discovery of highly potent human glutaminyl cyclase (QC) inhibitors as anti-Alzheimer&#039;s agents by the combination of pharmacophore-based and structure-based design European Journal of Medicinal Chemistry, 2021-09-08;226(0):113819. 2021-09-08 [PMID: 34536669] (Bioassay, Human)

Bioinformatics

• Gene Symbol: QPCT
• Uniprot:
  • Q16769()