Phage Display Peptide Library Related Issues-Special Topic on Phage Display Technology-Tekbiotech-Yeast Display Service,Phage display technology

Phage Display Peptide Library Related Issues

I. What are the different functions and uses of pIII and pVIII proteins in phage display technology?

Answer:

pIII and pVIII proteins play different roles in phage display technology. pIII protein is a minor coat protein on the surface of phage, mainly used to display foreign proteins and peptides. Its surface is easily accessible, so it is suitable for displaying ligands of target receptors, enzyme blockers, protein/DNA-protein interaction studies, screening cDNA expression, antibody epitope mapping, engineering human antibodies, etc. pVIII protein is the main coat protein on the surface of phage, and can also be used to display foreign proteins and peptides. However, compared with pIII, the length of peptides available for display on the surface of pVIII protein is shorter and can only accommodate short peptides (<10 amino acid residues). In addition, the display of pVIII protein will lead to a multivalent binding effect, making the protein or peptide selected from the pVIII display library have a lower affinity with the target. Therefore, pIII and pVIII have different functions and uses in phage display technology.

II. What are the advantages and limitations of phage display technology compared with other technologies?

Answer:

Advantages: 1. Phage display technology can construct a highly diverse phage library, making it possible to quickly isolate and identify specific protein ligands. 2. Simple, economical and fast: Phage display technology is simpler, economical and faster than traditional random screening methods. Through standard molecular biology methods, a large number of exogenous peptides or proteins expressed on the surface of phages can be quickly generated. 3. Structural analysis: The ligands screened by phage display technology can be structurally analyzed to understand the interaction between ligands and targets in more detail, providing more information for the next step of drug development and process development.

Limitations: 1. Phage display technology is a cell-based method that is limited by DNA conversion efficiency and the toxicity of displayed molecules to host cells. 2. Library size limitation: Due to the limitation of DNA conversion efficiency, the library size of phage display technology is limited. 3. Size and properties of displayed molecules: Phage display technology has certain limitations on the size and properties of displayed molecules. Proteins/peptides of different sizes and properties may need to be displayed using different types of phages.

III. What are the steps of in vivo screening and in vitro screening?

Answer:

Bioscreening is the most commonly used in vitro screening method for identifying and isolating ligands that bind to a target. Bioscreening involves the following steps: (1) Target immobilization: The purified target of interest is immobilized on a plate, which can also be performed on adherent cells (the desired target is a cell surface receptor); (2) Phage binding: Add a phage library and bind to the target under conditions suitable for binding; (3) Washing: Unbound phages are removed; (4) Phage elution: Due to the high stability of filamentous phages, bound phages can be eluted using a variety of methods. A common method for recovering bound phages is to disrupt the interaction between the displayed ligand and the target by changing the pH or adding a competing ligand, denaturant, or protease (for example, a protease cleavage site is involved between the displayed protein/peptide at the N-terminus and the coat protein itself); (5) Increase stringency: The eluted phages are then amplified in bacterial cells and repeated for several rounds of bioscreening (usually 3-5 rounds). This tends to select for phages with low affinity/non-specific binding to the target of interest; (6) Identify the target peptide sequence using DNA sequencing.

In vivo screening can be used to identify aptamer ligands that can home to specific tissues or organs. For example, phages can be injected intravenously into an animal and allowed to circulate for a period of time. The phages are then recovered from the selected organ, amplified, and their DNA sequenced. In this way, "non-specific" phages tend to be distributed throughout the animal, while phages with more "selective" target ligands are concentrated in specific tissues. Phage-derived ligands are specific for organs or tissues and may be used as diagnostic tools or disease treatments by coupling the phages to drugs or assembling the phages on drug-containing nanoparticles.

Based on the phage display platform, TekBiotech can construct random 7-peptide libraries, 12-peptide libraries, cyclic 7-peptide libraries, etc. It can not only provide customers with a variety of linear peptide library construction services based on T7 phage and M13 phage, but also provide customers with high-quality cyclic peptide phage display library construction and screening services.

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