Random combinatorial library

There are two general methods for identifying antibody epitopes (1) evaluate peptides whose composition is based on the sequence of the native protein, or (2) evaluate peptides that are selected from a random combinatorial peptide library. The former is only effective if the epitope is composed of a linear sequence of amino acids in the native protein sequence. If it is, then analysis of overlapping peptides from the native protein is a simple method for identifying antibody epitopes. Each peptide is tested for immunoreactivity to the antibody. Those peptides that are immunoreactive contain the epitope. The other method of epitope identihcation, selection from a random combinatorial library, will identify peptides that represent both linear and conformationally dependent epitopes. The drawback of this method is that biopanning from a random combinatorial peptide library is more time consuming. Phage-displayed peptide libraries have previously been used for epitope identification in this context. ... 

By contrast, peptide 124 was found to populate a canonical type 11 p-tum in its lowest energy ensemble (Fig. 13b). That it was discovered from a random combinatorial library speaks to the possibly privileged nature of this secondary structure element [188]. Furthermore [183], the structure validated a number of hypotheses based on sequence variations and truncation studies that support a remote hydroxyl-directed mechanism of action for the oxidation of the 6,7-position of famesol by 124. Importantly, a sidechain ether is implicated as a key acceptor of a hydrogen bond from famesol. 

EMPl, selected by phage display from random peptide libraries, demonstrates that a dimer of a 20-residue peptide can mimic the function of a monomeric 166-residue protein. In contrast to the minimized Z domain, this selected peptide shares neither the sequence nor the structure of the natural hormone. Thus, there can be a number of ways to solve a molecular recognition problem, and combinatorial methods such as phage display allow us to sort through a multitude of structural scaffolds to discover novel solutions. 

Combinatorial chemistry, a new chapter of organic synthesis, is now developing rapidly. This new approach to synthesizing large designed or random chemical libraries through application of solid phase synthetic methods, promises to revolutionize the process of drug discovery in the pharmaceutical industry.24... 

In general, reagent-based selection is much faster and more convenient to execute in the laboratory as compared with the product-based selection. On the other hand, the latter strategy usually provides more accurate results. There exists a potential to combine both approaches to achieve more optimal results, particularly in the case of large exploratory virtual combinatorial libraries, for which mass random synthesis and screening are not economically feasible. In this article, we demonstrated the usefulness of property-based approach for selection of optimal GPCR ligands. 

Corbett, P. T Sanders, J. K. M. Otto, S. Systems chemistry Pattern formation in random dynamic combinatorial libraries. Angew. Chem. Int. Ed. 2007, 46, 8858-8861. 

The principal aim of molecular diversity analysis is to identify structurally diverse (synonyms are dissimilar, disparate, and heterogeneous) sets of compounds that can then be tested for bioactivity, the assumption being that a structurally diverse set will generate more structure-activity information than will a set of compounds identified at random. The sets of compounds can be selected from an existing corporate or public database, or can be the result of a systematic combinatorial library design process (4,5). 

Generate a trial solution to the underlying problem. For combinatorial library design, a random selection of a subset of building blocks is generated. 

Combinatorial libraries are limited by the number of sequences that can be synthesized. For example, a library consisting of one molecule each of a 60-nudeotide sequence randomized at each position, would have a mass of >1014 g, well beyond the capacity for synthesis and manipulation. Thus, even if nucleotide addition is random at all the steps during synthesis of the oligonudeotide only a minority of the sequences can be present in the output from a laboratory-scale chemical DNA synthesis reaction. In analyzing these random but incomplete libraries, the protocol is efficient enough to allow selection of aptamers of lowest dissociation constants (Kd) from the mixture after a small number of repetitive selection and amplification cycles. Once a smaller population of oligonucleotides is amplified, the aptamer sequences can be used as the basis for constructing a less complex library for further selection. 

Kang, A S, Jones, T M, and Burton, D R. (1991) Antibody redesign by chain shuffling from random combinatorial immunoglobulin libraries Proc Natl Acad Sci USA 88, 11,120-11,123... 

In 1991, we first introduced the one-bead one-compound (OBOC ) combinatorial library method.1 Since then, it has been successfully applied to the identification of ligands for a large number of biological targets.2,3 Using well-established on-bead binding or functional assays, the OBOC method is highly efficient and practical. A random library of millions of beads can be rapidly screened in parallel for a specific acceptor molecule (receptor, antibody, enzyme, virus, etc.). The amount of acceptor needed is minute compared to solution phase assay in microtiter plates. The positive beads with active compounds are easily isolated and subjected to structural determination. For peptides that contain natural amino acids and have a free N-terminus, we routinely use an automatic protein sequencer with Edman chemistry, which converts each a-amino acid sequentially to its phenylthiohydantoin (PTH) derivatives, to determine the structure of peptide on the positive beads. 

A variation of the pharmacophore library was seen in an approach used to determine CD4+ T cell epitopes.31 In a 14-mer peptide, the researchers keep residues in positions 1, 4, and 6 constant since they functioned as anchor positions for binding receptor, and proceeded to vary the other residues in the peptide. This allowed the use of a shorter synthetic route compared to one that would be needed if all positions were to be varied. Limiting the size of the library allowed the authors to obtain better assays. By doing a partial release of the peptides, the authors were able to find the final peptide that represented the epitope of the T cell receptor. In another example, the epitope to inhibit stimulation of the thyrotropin receptor also was found via combinatorial libraries.32 Since the synthesis of a totally random hexapeptide library was deemed impractical, the authors opted to hold one position constant while the other five residues were randomized. This method was repeated for each residue in the peptide. The residues that were determined to be the most active were used as a basis for a second-generation library. The only limitation of the library was not the quantity of product synthesized, but to properly pinpoint the peptides in an assay. 

Another possibility to screen for active peptides without prior knowledge of a starting sequence is use of a random peptide library. This array contains randomized peptide sequences (61). Compared with the combinatorial peptide... 

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