Peptide nanofibrils (PNF) in retroviral gene transfer
Self-assembly mechanisms of PNFs
Recent advances in molecular medicine have greatly simplified the transfer of genetic material into cells for a myriad of applications, such as the development of mRNA vaccines and gene therapy. Gene therapy approaches can be used to treat diseases that are both inherited, such as those caused by gene defects, and acquired diseases, such as cancer. Among others, retroviral gene transfer is the most commonly used method of choice to introduce genetic material into cellular genome. However, the efficiency of retroviral gene transfer is still limited. For most applications, low concentrations of viral vectors must be used to avoid toxicity in the living cells and consequently the efficiency of the viral vectors in attaching to the cells is lowered. In order to overcome these challenges in retroviral gene transfer, researchers strive to develop new synthetic materials, composed of polymers, peptides and lipids, that will enhance gene transfer and are nontoxic, biodegradable and sustainable. One of these biomaterials are self-assembling peptide nanofibrils (PNFs), which are supramolecular amyloid-like nanofibers popular due to their high stability, resistance to degradation, biocompatibility and dynamic properties that can be tuned with external stimuli.
To address these challenges, the proposed research project will focus on development of physics-based and data-driven models and simulation strategies to investigate the relationship between structure, morphology, and materials properties of PNFs for enhancement of efficiencies of retroviral gene transfer. Using the models and methods developed, this project also aims to answer questions about dynamic properties and self-assembly pathways of supramolecular synthetic and naturally-occuring amyloid-like nanofibrils. Ultimately, we aim to advance the field further and utilize computational methods not only for understanding chemical and physical processes in soft matter, but also develop new chemical and physical concepts using soft matter as a tool in a smart and rational manner. The insights gained in this project about structure-property relationships of amyloid-like nanofibers will be used to design new materials that would give the best performance, reduce the need for time-consuming trial and error experiments in laboratories by evaluating a large number of different peptide sequences and identifying possible candidates to be synthesized by our experimental partners. This knowledge will be extremely beneficial to the pharmaceutical community that is aimed at development of therapeutics to shift the equilibrium away from formation of fibers and stabilize the monomeric forms of amyloid forming proteins, such as Aβ and tau proteins.
References
2024
Multiscale Simulations of Self-Assembling Peptides: Surface and Core Hydrophobicity Determine Fibril Stability and Amyloid Aggregation
Aysenur Iscen, Kübra Kaygisiz, Christopher V. Synatschke, and 2 more authors
Assemblies of peptides and proteins through specific intermolecular interactions set the basis for macroscopic materials found in nature. Peptides provide easily tunable hydrogen-bonding interactions, which can lead to the formation of ordered structures such as highly stable β-sheets that can form amyloid-like supramolecular peptide nanofibrils (PNFs). PNFs are of special interest, as they could be considered as mimics of various fibrillar structures found in nature. In their ability to serve as supramolecular scaffolds, they could mimic certain features of the extracellular matrix to provide stability, interact with pathogens such as virions, and transduce signals between the outside and inside of cells. Many PNFs have been reported that reveal rich bioactivities. PNFs supporting neuronal cell growth or lentiviral gene transduction have been studied systematically, and their material properties were correlated to bioactivities. However, the impact of the structure of PNFs, their dynamics, and stabilities on their unique functions is still elusive. Herein, we provide a microscopic view of the self-assembled PNFs to unravel how the amino acid sequence of self-assembling peptides affects their secondary structure and dynamic properties of the peptides within supramolecular fibrils. Based on sequence truncation, amino acid substitution, and sequence reordering, we demonstrate that peptide–peptide aggregation propensity is critical to form bioactive β-sheet-rich structures. In contrast to previous studies, a very high peptide aggregation propensity reduces bioactivity due to intermolecular misalignment and instabilities that emerge when fibrils are in close proximity to other fibrils in solution. Our multiscale simulation approach correlates changes in biological activity back to single amino acid modifications. Understanding these relationships could lead to future material discoveries where the molecular sequence predictably determines the macroscopic properties and biological activity. In addition, our studies may provide new insights into naturally occurring amyloid fibrils in neurodegenerative diseases.
2023
Data-mining unveils structure–property–activity correlation of viral infectivity enhancing self-assembling peptides
Kübra Kaygisiz, Lena Rauch-Wirth, Arghya Dutta, and 6 more authors
Gene therapy via retroviral vectors holds great promise for treating a variety of serious diseases. It requires the use of additives to boost infectivity. Amyloid-like peptide nanofibers (PNFs) were shown to efficiently enhance retroviral gene transfer. However, the underlying mode of action of these peptides remains largely unknown. Data-mining is an efficient method to systematically study structure–function relationship and unveil patterns in a database. This data-mining study elucidates the multi-scale structure–property–activity relationship of transduction enhancing peptides for retroviral gene transfer. In contrast to previous reports, we find that not the amyloid fibrils themselves, but rather µm-sized β-sheet rich aggregates enhance infectivity. Specifically, microscopic aggregation of β-sheet rich amyloid structures with a hydrophobic surface pattern and positive surface charge are identified as key material properties. We validate the reliability of the amphiphilic sequence pattern and the general applicability of the key properties by rationally creating new active sequences and identifying short amyloidal peptides from various pathogenic and functional origin. Data-mining—even for small datasets—enables the development of new efficient retroviral transduction enhancers and provides important insights into the diverse bioactivity of the functional material class of amyloids.
2021
Supramolecular Peptide Nanofibrils with Optimized Sequences and Molecular Structures for Efficient Retroviral Transduction
Stefanie Sieste, Thomas Mack, Edina Lump, and 11 more authors
Abstract Amyloid-like peptide nanofibrils (PNFs) are abundant in nature providing rich bioactivities and playing both functional and pathological roles. The structural features responsible for their unique bioactivities are, however, still elusive. Supramolecular nanostructures are notoriously challenging to optimize, as sequence changes affect self-assembly, fibril morphologies, and biorecognition. Herein, the first sequence optimization of PNFs, derived from the peptide enhancing factor-C (EF-C, QCKIKQIINMWQ), for enhanced retroviral gene transduction via a multiparameter and a multiscale approach is reported. Retroviral gene transfer is the method of choice for the stable delivery of genetic information into cells offering great perspectives for the treatment of genetic disorders. Single fibril imaging, zeta potential, vibrational spectroscopy, and quantitative retroviral transduction assays provide the structure parameters responsible for PNF assembly, fibrils morphology, secondary and quaternary structure, and PNF-virus-cell interactions. Optimized peptide sequences such as the 7-mer, CKFKFQF, have been obtained quantitatively forming supramolecular nanofibrils with high intermolecular β-sheet content that efficiently bind virions and attach to cellular membranes revealing efficient retroviral gene transfer.
2013
Peptide nanofibrils boost retroviral gene transfer and provide a rapid means for concentrating viruses
Maral Yolamanova, Christoph Meier, Alexey K. Shaytan, and 26 more authors
Inefficient gene transfer and low virion concentrations are common limitations of retroviral transduction1. We and others have previously shown that peptides derived from human semen form amyloid fibrils that boost retroviral gene delivery by promoting virion attachment to the target cells2,3,4,5,6,7,8. However, application of these natural fibril-forming peptides is limited by moderate efficiencies, the high costs of peptide synthesis, and variability in fibril size and formation kinetics. Here, we report the development of nanofibrils that self-assemble in aqueous solution from a 12-residue peptide, termed enhancing factor C (EF-C). These artificial nanofibrils enhance retroviral gene transfer substantially more efficiently than semen-derived fibrils or other transduction enhancers. Moreover, EF-C nanofibrils allow the concentration of retroviral vectors by conventional low-speed centrifugation, and are safe and effective, as assessed in an ex vivo gene transfer study. Our results show that EF-C fibrils comprise a highly versatile, convenient and broadly applicable nanomaterial that holds the potential to significantly facilitate retroviral gene transfer in basic research and clinical applications.
