Engineering Alkaline-Stable Barley Stripe Mosaic Virus-Like Particles for Efficient Surface Modification.

Viruses and virus-like particles are powerful templates for materials synthesis because of their capacity for precise protein engineering and diverse surface functionalization. We recently developed a recombinant bacterial expression system for the production of barley stripe mosaic virus-like particles (BSMV VLPs). However, the applicability of this biotemplate was limited by low stability in alkaline conditions and a lack of chemical handles for ligand attachment. Here, we identify and validate novel residues in the BSMV Caspar carboxylate clusters that mediate virion disassembly through repulsive interactions at high pH. Point mutations of these residues to create attractive interactions that increase rod length ~2 fold, with an average rod length of 91 nm under alkaline conditions. To enable diverse chemical surface functionalization, we also introduce reactive lysine residues at the C-terminus of BSMV coat protein, which is presented on the VLP surface. Chemical conjugation reactions with this lysine proceed more quickly under alkaline conditions. Thus, our alkaline-stable VLP mutants are more suitable for rapid surface functionalization of long nanorods. This work validates novel residues involved in BSMV VLP assembly and demonstrates the feasibility of chemical functionalization of BSMV VLPs for the first time, enabling novel biomedical and chemical applications.

Surface Functionalization of Rod-Shaped Viral Particles for Biomedical Applications.

While synthetic nanoparticles play a very important role in modern medicine, concerns regarding toxicity, sustainability, stability, and dispersity are drawing increasing attention to naturally derived alternatives. Rod-shaped plant viruses and virus-like particles (VLPs) are biological nanoparticles with powerful advantages such as biocompatibility, tunable size and aspect ratio, monodispersity, and multivalency. These properties facilitate controlled biodistribution and tissue targeting for powerful applications in medicine. Ongoing research efforts focus on functionalizing or otherwise engineering these structures for a myriad of applications, including vaccines, imaging, and drug delivery. These include chemical and biological strategies for conjugation to small molecule chemical dyes, drugs, metals, polymers, peptides, proteins, carbohydrates, and nucleic acids. Many strategies are available and vary greatly in efficiency, modularity, selectivity, and simplicity. This review provides a comprehensive summary of VLP functionalization approaches while highlighting biomedically relevant examples. Limitations of current strategies and opportunities for further advancement will also be discussed.

ATDC binds to KEAP1 to drive NRF2-mediated tumorigenesis and chemoresistance in pancreatic cancer.

Pancreatic ductal adenocarcinoma is a lethal disease characterized by late diagnosis, propensity for early metastasis and resistance to chemotherapy. Little is known about the mechanisms that drive innate therapeutic resistance in pancreatic cancer. The ataxia-telangiectasia group D-associated gene (ATDC) is overexpressed in pancreatic cancer and promotes tumor growth and metastasis. Our study reveals that increased ATDC levels protect cancer cells from reactive oxygen species (ROS) via stabilization of nuclear factor erythroid 2-related factor 2 (NRF2). Mechanistically, ATDC binds to Kelch-like ECH-associated protein 1 (KEAP1), the principal regulator of NRF2 degradation, and thereby prevents degradation of NRF2 resulting in activation of a NRF2-dependent transcriptional program, reduced intracellular ROS and enhanced chemoresistance. Our findings define a novel role of ATDC in regulating redox balance and chemotherapeutic resistance by modulating NRF2 activity.

Decomplexation as a rate limitation in the thiol-Michael addition of N-acrylamides.

The thiol-Michael addition is a popular, selective, high-yield "click" reaction utilized for applications ranging from small-molecule synthesis to polymer or surface modification. Here, we combined experimental and quantum mechanical modeling approaches using density functional theory (DFT) to examine the thiol-Michael reaction of N-allyl-N-acrylamide monomers used to prepare sequence-defined oligothioetheramides (oligoTEAs). Experimentally, the reaction was evaluated with two fluorous tagged thiols and several monomers at room temperature (22 °C and 40 °C). Using the Eyring equation, the activation energies (enthalpies) were calculated, observing a wide range of energy barriers ranging from 28 kJ mol-1 to 108 kJ mol-1 within the same alkene class. Computationally, DFT coupled with the Nudged Elastic Band method was used to calculate the entire reaction coordinate of each monomer reaction using the B97-D3 functional and a hybrid implicit-explicit methanol solvation approach. The thiol-Michael reaction is traditionally rate-limited by the propagation or chain-transfer steps. However, our test case with N-acrylamides and fluorous thiols revealed experimental and computational data produced satisfactory agreement only when we considered a previously unconsidered step that we termed "product decomplexation", which occurs as the product physically dissociates from other co-reactants after chain transfer. Five monomers were investigated to support this finding, capturing a range of functional groups varying in alkyl chain length (methyl to hexyl) and aromaticity (benzyl and ethylenephenyl). Increased substrate alkyl chain length increased activation energy, explained by the inductive effect. Aromatic ring-stacking configurations significantly impacted the activation energy and contributed to improved molecular packing density. Hydrogen-bonding between reactants increased the activation energy emphasizing the rate-limitation of the product decomplexation. Our findings begin to describe a new structure-kinetic relationship for thiol-Michael acceptors to enable further design of reactive monomers for synthetic polymers and biomaterials.