Analysis the Extracellular Loop II of κ-Opioid Receptor (196–228) with Opioid Peptide Dynorphin Interaction by Quartz Crystal Microbalance (QCM) / -Dissipation (QCM-D) (CAT#: STEM-MB-0742-CJ)

Introduction

κ-Opioid receptor is a member of the opioid receptor family and selectively interacts with the opioid peptide dynorphin. Extracellular loop II (ECL-II) of the κ-opioid receptor displays an amphiphilic helix in membrane environments and the N-terminal α-helix of dynorphin A(1–17) is inserted into the membrane with the tilt angle of 21° to the bilayer normal. ECL-II peptides (1–33), corresponding to 196–228 of κ-opioid receptor with [1-13C]- or [3-13C]-labeled amino acids were incorporated into large [dimyristoylphosphatidyl choline (DMPC)/ dihexanoylphosphatidyl choline (DHPC) = 3, q = 3] and small bicelle (q = 1) systems.

Dynorphin is a neuropeptide involved in pain, addiction and mood regulation. It exerts its activity by binding to the kappa opioid receptor (KOP) which belongs to the large family of G protein-coupled receptors. The dynorphin peptide was discovered in 1975, while its receptor was cloned in 1993.

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Principle Applications Procedure Materials Notes Related Products

Principle

The basic principle of QCM is the piezoelectric effect of quartz crystal. Upon an alternating voltage applied to the two poles of the quartz crystal, the quartz crystal of QCM produces mechanical vibration or oscillation. This resonance is greatly sensitive to the thickness of the crystal/electrode system and the frequency of the acoustic frequency. Once the mass on the quartz crystal increases or decreases, the resonance will be disturbed and the resonance frequency of the quartz crystal will change. Thus, the mass variation on the quartz crystal electrode surface is transformed into the frequency variation of the output electric signal of the quartz crystal oscillation and is further transformed into the high-precision data by the computer.

Applications

Biochemistry; Medical; Pharmacology

Procedure

1. Sensor preparation, reduce the risk of damaging or contaminating the sensor surface.
2. Preparation of sample liquids.
3. Places the sensor into the flow cell and ensures perfect alignment.
4. Instant identification of specific sensors by unique ID number.
5. Using quarter turn bayonet type sealed flow cell.
6. Enter the experiment details in the information panel.
7. Display frequency and dissipation in real time.
8. Observe the bubble-free filling through the transparent window.
9. Electronic log file detects all steps in the measurement process.
10. Export and analyze the data.

Materials

• Sample: Proteins; Microbes; Nucleic Acids; Enzymes; Cells; Polymers; Small molecules; Antibody; Lipids; Virus-like particles (VLPs);
• Equipment: Quartz Crystal Microbalance (QCM) / Quartz Crystal Microbalance-Dissipation (QCM-D)
• Running buffer
• (Optional) PBS solution

Notes

1. QCM is an extremely sensitive mass balance that measures nanogram to microgram level changes in mass per unit area.
2. QCM detection limit is approximately 250 ng mL−1, which is similar to that of the existing enzyme-linked immunosorbent assay (ELISA) method.
3. Unlike traditional QCM, the QCM with an added D measures an additional parameter, namely the dissipation factor. so QCM-D has great advantages in providing binding kinetics information by virtue of its label-free breadth of applicability, low cost, and sensing unique mass distribution and viscoelastic behavior aspects of whole-cell responses in real time.
4. QCM-D was able to measure nanoparticle-induced platelet microaggregation for all nanoparticles tested at concentrations that were undetectable by light aggregometry and flow cytometry.

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