PNA Hybrid Sequences as Recognition Units in SNARE‐Protein‐Mimicking Peptides

Abstract

Membrane fusion is an essential process in nature and is often accomplished by the specific interaction of SNARE proteins. SNARE model systems, in which SNARE domains are replaced by small artificial units, represent valuable tools to study membrane fusion in vitro. The synthesis and analysis is presented of SNARE model peptides that exhibit a recognition motif composed of two different types of peptide nucleic acid (PNA) sequences. This novel recognition unit is designed to mimic the SNARE zippering mechanism that initiates SNARE-mediated fusion. It contains N-(2-aminoethyl)glycine-PNA (aeg-PNA) and alanyl-PNA, which both recognize the respective complementary strand but differ in duplex topology and duplex formation kinetics. The duplex formation of PNA hybrid oligomers as well as the fusogenicity of the model peptides in lipid-mixing assays were characterized and the peptides were found to induce liposome fusion. As an unexpected discovery, peptides with a recognition unit containing only five aeg-PNA nucleo amino acids were sufficient and most efficient to induce liposome fusion.

Neuronal SNARE complex assembly guided by Munc18‐1 and Munc13‐1

“…Third, the fusion kinetics obtained from in vitro reconstitution assay using the SNAREs alone is not comparable to that observed in vivo . In addition, previous studies reported that two complementary paired nucleic acid strands linked by the TMRs of syntaxin‐1 and synaptobrevin‐2 could mediate in vitro liposome fusion as well [ 65 , 66 ]. Another study revealed that artificial coiled‐coil peptides linked by the JLRs and TMRs of the SNAREs could induce liposome fusion[ 67 ].…”

Membrane fusion mediated by peptidic SNARE protein analogues: Evaluation of FRET‐based bulk leaflet mixing assays

“…In the present study, different conditions for bulk leaflet mixing assays performed with established SNARE‐like model peptides are inspected and evaluated. The model peptides either contain peptide nucleic acid (PNA) 6 or coiled‐coil forming peptide segments (E3/K3) 7 as recognition units (Table 1). Both structural motifs have already been used in earlier studies to explore membrane fusion 8,9 .…”

“…The preparation of liposomes as well as the bulk leaflet mixing assays were performed as described earlier. 6,7 In brief, solutions of the respective lipids and peptides were mixed and the organic solvents were evaporated to obtain lipid films. The lipid films were rehydrated in buffer and proteoliposomes were formed by extrusion.…”

Prebiotic Peptides: Molecular Hubs in the Origin of Life

“…Beginning in the mid-1990s, scientists have explored the properties of a class of PNAs termed alanyl nucleic acids (ANAs). − As opposed to dipeptide-based PNA structures, in which a nucleobase is attached to every other amino acid to match the six-atom backbone repeat geometry found in RNA and DNA, ANA contains a nucleobase-substituted amino acid at every position in the sequence (Chart ). Amino acids are present in ANA with alternating chiral configurations.…”