Genetically Encoded Libraries of Constrained Peptides.

Many therapeutic peptides can still be improved with respect to target specificity, target affinity, resistance to peptidases/proteases, physical stability, and capacity to pass through membranes required for oral delivery. Several modifications can improve the peptides' properties, in particular those that impose (a) conformational constraint(s). Screening of constrained peptides and the identification of hits is greatly facilitated by the generation of genetically encoded libraries.

Phage display and selection of lanthipeptides on the carboxy-terminus of the gene-3 minor coat protein.

Ribosomally synthesized and post-translationally modified peptides (RiPPs) are an emerging class of natural products with drug-like properties. To fully exploit the potential of RiPPs as peptide drug candidates, tools for their systematic engineering are required. Here we report the engineering of lanthipeptides, a subclass of RiPPs characterized by multiple thioether cycles that are enzymatically introduced in a regio- and stereospecific manner, by phage display.

Binding proteins internalized by PTD-fused ligands allow the intracellular sequestration of selected targets by ligand exchange.

The targeted inactivation of intracellular molecules has important therapeutic potential. For this purpose, it could be envisioned to introduce specifically designed binding proteins into cells by covalent linkage to protein transduction domains (PTDs). However, stable linkage of a PTD to a cargo may affect its conformation and, hence, its binding property inside the cell.

Transtactin: a universal transmembrane delivery system for Strep-tag II-fused cargos.

The delivery of molecules into cells poses a critical problem that has to be solved for the development of diagnostic tools and therapeutic agents acting on intracellular targets. Cargos which by themselves cannot penetrate cellular membranes due to their biophysical properties can achieve cell membrane permeability by fusion to protein transduction domains (PTDs). Here, we engineered a universal delivery system based on PTD-fused Strep-Tactin, which we named Transtactin.

Membrane-permeable cygnets: rapid cellular internalization of fluorescent cGMP-indicators.

We have recently developed genetically encoded cGMP-indicators (cygnets) which have enabled us to study the spatial and temporal dynamics of intracellular cGMP in single cultured cells (1). However, primary mammalian cell types (dissociated cells or acute tissue samples) are often difficult to maintain undifferentiated in culture and the current established methods of introducing molecular reporters in single cells are laborious (micro-injection) and/or require cell culture techniques to accommodate the 1-2 day lag time of genetically mediated reporter expression. Here, we present an alternative, non-genetic method to rapidly introduce cGMP-indicators into cells and intact tissues using membrane permeable peptides (MPP).