Peptide Library Services
Proteins are large molecules and are usually ineffective as drugs due to delivery and stability issues. As a large molecule, a single protein may have multiple biological functions where each function is defined by localized interactions of a specific sequence of amino acids in the protein with another protein or a non-protein ligand. In the peptide-based drug discovery process, the conventional one protein-one experiment strategy is too time-consuming and costly. What is required is high throughput screening that screens a large number of compounds quickly and efficiently in a parallel manner.
Drug development is broken down into several distinct steps. The first step is to "map" the active sequences (i.e. epitope) of a protein, a process called epitope mapping that will determine the minimum sequence of a peptide that makes up the active domain of the protein. Epitope mapping synthesizes a peptide library that consists of overlapping peptide sequences that makes up the original protein. To determine the sequences of a peptide library, the selection process has the following considerations:
- As the length of the peptide grows, the number of peptides to synthesize decreases.
- As the offset number grows (i.e. the number of residues that the peptide sequence shifts from the original protein sequence), the number of peptides to synthesize decreases.
- As the peptide sequence grows longer, the potential to achieve multiple hits (i.e. the peptide sequences that contain all of the essential residues in the epitope) increases.
The length of the protein sequence will determine the number of peptides in the library. Choosing a longer sequence and a shorter offset number would be ideal, but the monetary cost could become too great. Shorter peptide lengths, in contrast, will lead to more peptide sequences to synthesize and are more economical. Because it may be difficult to predetermine the ideal minimum number of peptides needed, the common practice is to use 8 to 20 residues (preferably in the 12 to 16 range) with the offset number being approximately 1/3 of the peptide length.
Peptide Library Types and Strategies
The next step in drug development is peptide sequence optimization, the step that will determine the structure and functional relationships of the targeted epitope, utilizing four practical strategies.
- Alanine Scanning Library: Alanine (chosen because it is the smallest amino acid that maintains chirality) is systematically substituted for each amino acid position in the epitope. If alanine takes the place of an essential amino acid, the result would be a significant reduction in activity. The relative importance of that specific amino acid could also be measured by the degree of activity reduction.
- Truncation Library: Involves the systematic truncation of the flanking residues to determine the minimum length required for optimum peptide activity.
- Random Library: A shotgun approach. A mixture of all 20 amino acids, or a set combination of amino acids, is simultaneously substituted for selected residues in the peptide sequence (i.e. the wobble sequence). The mixtures of these random libraries are then analyzed.
- Positional Scanning Library: A selected position or positions in a peptide sequence are each systematically replaced with different amino acids. The resulting change in activity reveals the preferred amino acid residues at these positions.
The last step for peptide-based drug development is sequence stabilization. A structural stabilization of the peptide needs to be done to preserve their potency over time. Three different strategies can be employed to achieve this goal:
- The most common method is to substitute selected amino acids with non-standard amino acids, like either homologs of natural amino acids (ex. ornithine, homolysine, norleucine, and norvaline) or the chiral analogs (D-forms) or the naturally-occuring amino acids (L-forms).
- Another method is to incorporate intramolecular bridges to form cyclic structures.
- The stabilization can be achieved through the chemical modification of the N- and C-termini (usually by acetylation and amidation, respectively).
Peptide Library & Epitope Mapping Citations
1. Karlsson, Erik A., et al. "Obesity Outweighs Protection Conferred by Adjuvanted Influenza Vaccination." mBio 7.4 (2016): e01144-16.Learn More »
2. Stair, Jacqueline L., et al. "Quantitative determination of single-bead metal content from a peptide combinatorial library." Journal of Combinatorial Chemistry 8.6 (2006): 929-934.Learn More »
3. Falta, Michael T., et al. "Identification of beryllium-dependent peptides recognized by CD4+ T cells in chronic beryllium disease." The Journal of Experimental Medicine 210.7 (2013): 1403-1418.Learn More »
4. Ren, Yin, and Sangeeta N. Bhatia. "Targeted delivery of nucleic acids." U.S. Patent No. 9,006,415. 14 Apr. 2015.Learn More »
5. Wang, Xuelian, et al. "Memory T cells specific for novel human papillomavirus type 16 (HPV16) E6 epitopes in women whose HPV16 infection has become undetectable." Clinical and Vaccine Immunology 15.6 (2008): 937-945.
Learn More »
6. Wang, Xuelian, et al. "A novel CD4 T-cell epitope described from one of the cervical cancer patients vaccinated with HPV 16 or 18 E7-pulsed dendritic cells." Cancer Immunology, Immunotherapy 58.2 (2009): 301-308.Learn More »
7. Shojaei, Farbod, et al. "Osteopontin induces growth of metastatic tumors in a preclinical model of non-small lung cancer." Journal of Experimental & Clinical Cancer Research 31.1 (2012): 1.Learn More »
PEPscreen® Peptide Library Products
CPC Scientific is very pleased to announce an agreement with Sigma-Aldrich, allowing CPC Scientific to sell PEPscreen® custom peptide libraries, effective immediately. PEPscreen® is an empowering technology for peptide based drug discovery and Sigma's proprietary peptide synthesis platform enables fast and efficient high-throughput parallel synthesis of milligram quantities of peptides.
PEPscreen® Product Specifications:
- Quantity: 0.5 to 2 mg
- Modifications available
- Format: dried in 96-tube rack format
- QC: MALDI-TOF MS performed on all peptides ( Electronic PDF files of QC data and peptide directory supplied on CD )
- C-Terminal acid or amide
- Dispatch: approximately 7 business days
- Minimum order size is 24 peptides
Why use the PEPscreen® platform?
Peptide libraries can be highly variable, differing in length, N-termini, C-termini, the positions of the amino acid mixtures (x20), and the choices of non-standard amino acids. The PEPscreen® platform is the first truly flexible system to meet these requirements in a single run, fulfilling the need for a system that has high throughput screening (HTS) in a parallel manner. The PEPscreen® system is highly automated and flexible allowing sequences with lengths from 6 to 20 amino acids, N-termini with free amine oracetylation, C- termini with free acid or amidation, any mixture of commercially available amino acids for use in the random library optimization process, and the addition of any available non-standard amino acid. Chemical modifications are also available, including, phosphorylation, biotinylation, PEGylation, acylation and others, in addition to dye labeling with FIc, FITC, Dansyl, Dabcyl, Dabsyl, TAMRA, Lissamine, etc. Between 0.5 mg to 2 mg of each peptide is delivered as dry film in individual tubes with capped lids arranged in a standard 8 x 12 tube plate for compatibility with high throughput assays. Each tube is triple labeled with alpha-numeric, barcode and 2D barcode system. Each peptide passes both MS analysis and the final gross weight criteria before shipment. All MS data are burned onto a CD and supplied with the final product. HPLC analysis can also be performed for a reasonable fee by prior arrangement.
How are PEPscreen® handled and stored?
Peptides must be completely solubilized to be fully active. To dissolve PEPscreen peptide sets, the initial solubilization attempt must be accomplished with a stronger solvent, such as DMSO, DMF, acetic acid, etc., than that which would be used to dissolve individual peptides. This would be followed with a sonication of the peptide sample for several minutes. To further dissolve any remaining insoluble peptides after the first attempt, more benign buffers can be added to either the whole set or only to the insoluble peptides.
For further information and pricing, please contact us.
PEPscreen® is a registered trademark of Sigma-Aldrich Biotechnology, L.P. and Sigma-Aldrich Co.