Bacterial Display and Screening of Posttranslationally Thioether-Stabilized Peptides
| Autor: | Gert N. Moll, Anneke Kuipers, Rick Rink, Tjibbe Bosma, Louwe de Vries, Erna Bulten |
|---|---|
| Rok vydání: | 2011 |
| Předmět: |
Phage display
Peptide Sulfides Applied Microbiology and Biotechnology chemistry.chemical_compound Cell Wall Sortase Mass Screening mRNA display Enzymology and Protein Engineering chemistry.chemical_classification Bacterial display Ecology biology Protein Stability Lactococcus lactis biology.organism_classification Cyclic peptide Protein Transport chemistry Biochemistry Peptidoglycan Peptides Protein Processing Post-Translational Food Science Biotechnology |
| Zdroj: | Applied and Environmental Microbiology. 77:6794-6801 |
| ISSN: | 1098-5336 0099-2240 |
| Popis: | A major hurdle in the application of therapeutic peptides is their rapid degradation by peptidases. Thioether bridges effectively protect therapeutic peptides against breakdown, thereby strongly increasing bioavailability, enabling oral and pulmonary delivery and potentially significantly optimizing the receptor interaction of selected variants. To efficiently select optimal variants, a library of DNA-coupled thioether-bridged peptides is highly desirable. Here, we present a unique cell surface display system of thioether-bridged peptides and successfully demonstrate highly selective screening. Peptides are posttranslationally modified by thioether bridge-installing enzymes in Lactococcus lactis, followed by export and sortase-mediated covalent coupling to the lactococcal cell wall. This allows the combinatorial optimization and selection of medically and economically highly important therapeutic peptides with strongly enhanced therapeutic potential. Surface display of peptides is a powerful technology for selecting peptides with desired properties from combinatorial libraries. A peptide display system comprises a scaffold—e.g., phage (39), bacterium (10), ribosome (25), or the yeast Saccharomyces cerevisiae (4)—for the displayed peptide and a link between the displayed peptide and the encoding DNA. Selection involves several rounds of target binding, washing, elution of specific binders, and then amplification of the selected peptides. Each round results in increasing enrichment of binders over the nonbinders. To date phage display is the best known display system. Bacterial cell surface display has been developed for Gramnegative and Gram-positive bacteria (22). In the latter case, cell surface display relies on the translational fusion of a peptide to an LPXTG cell wall-anchoring motif such as that of the Lactococcus lactis PrtP protease (15, 38). This anchoring mechanism requires processing by a sortase for covalent anchoring of the peptide to the peptidoglycan of the bacterial cell wall (36). In this way the peptide and the encoding DNA are linked allowing selection, followed by rapid identification. Various peptide display libraries have been designed, including linear peptides, disulfide-linked cyclic peptides (40), and chemically modified peptides (1), for instance, peptides coupled to an organic core (11) and peptides with nonnatural amino acids (8). This has resulted in the identification of various useful peptides, including therapeutically effective peptides (20). In nature, the majority of therapeutic (poly)peptides bears some form of posttranslational modification (PTM) which modulates the peptide’s function (41). Combining display technology and true posttranslational modification of pep |
| Databáze: | OpenAIRE |
| Externí odkaz: |
|