Quartz-Crystal Microbalance Biosensor-Based Biopanning to Study Protein-Drug Interactions
Published: June 29, 2023
Abstract
Source: Takakusagi, Y. Biosensor-based High Throughput Biopanning and Bioinformatics Analysis Strategy for the Global Validation of Drug-protein Interactions. J. Vis. Exp. (2020)
This video demonstrates a method to study protein-drug interactions using QCM biosensor-based biopanning of the T7 phage-displayed peptide library. The biopanning of phages displaying a specific peptide on the QCM biosensor allows specific isolation of high-affinity peptides, followed by amino acid sequencing, which helps to identify specific interaction sites on the target protein.
Protocol
NOTE: The following are the steps for screening drug-recognizing T7 phages using a QCM biosensor and recovering the screened phages via E. coli (BLT5615) infection. The protocols for the synthesis of a derivative of a small molecule that forms a self-assembled monolayer (SAM) and for the construction of the T7 phage-displayed 15-mer random peptide library can be found elsewhere. 1. Preparation of the QCM sensor chip Attach a ceramic sensor…
Representative Results
Figure 1: Schematic representation of the QCM biosensor-based biopanning of the T7 phage-displayed peptide library. A T7 phage library that displays random peptides is injected into the cuvette containing the buffer (under stirring) where the QCM biosensor chip is immersed and the frequency is stabilized. After monitoring the frequency reduction due to the binding of T7 phages to…
Offenlegungen
The authors have nothing to disclose.
Materials
| AFFINIXQN | ULVAC, Inc. (Tokyo, Japan) | QCM2008-STKIT | Contains Glass cuvette, stir magnet, operation and analysis software with a Windows PC |
| Ceramic Sensor Chip | ULVAC, Inc. (Tokyo, Japan) | QCMST27C | 4 sensor chips/package |
| Dimethyl sulfoxide | Sigma-Aldrich (St. Louis, MO, USA) |
D8418 | |
| Ethanol | Merck (Kenilworth, NJ, USA) | 09-0850 | |
| Immobilization kit for AFFINIX | ULVAC, Inc. (Tokyo, Japan) | QCMIMKT | SAM reagent and amine-coupling reagent |
| Liquid LB medium | Sigma-Aldrich (St. Louis, MO, USA) |
L3522 | Autoclave for 20 min |
| NaCl | Merck (Kenilworth, NJ, USA) | S3014 | |
| Tris | Merck (Kenilworth, NJ, USA) | 252859 |
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Quartz-Crystal Microbalance Biosensor-Based Biopanning to Study Protein-Drug Interactions. J. Vis. Exp. (Pending Publication), e21447, doi: (2023).