Biology subjects

Rudra, J. S.

Publications and source records attributed to Rudra, J. S..

5 recordsLinked to original sources

bioRxiv2025

Substituent-based Modulation of Self-Assembly and Immunogenicity of Amphipathic Peptides

Peptide-based biomaterials assembled through monomer-by-monomer self-assembly provide versatile platforms for biomedical applications due to their adjustable physicochemical properties, biocompatibility, and dynamic nature. The self-assembly process largely depends on primary sequence features, such as hydrophobicity, length, and charge, which influence the formation of various nanostructures, including fibrils and hydrogels. Amphipathic peptides, characterized by alternating polar and hydrophobic residues, are especially effective in forming supramolecular nanofibers stabilized by {pi}-{pi} interactions and hydrogen bonds. Chemical modifications, particularly on aromatic side chains, have proven to be a promising approach for controlling assembly morphology, stability, and biological activity. In organic chemistry, the use of chemical substituents, such as halogens, alkyl groups, or electron-donating and electron-withdrawing groups, has been widely employed to alter reactivity, stability, and molecular interactions for diverse applications, including catalysts, pharmaceuticals, and materials science. However, the influence of these substituents on peptide packing and in vivo immunogenicity remains relatively unexplored. In this study, we systematically examine how changes in the position and nature of substituents on benzyl groups attached to short amphipathic peptides affect self-assembly, fibril morphology, and immune responses. By introducing different electron-donating and withdrawing groups at the para-position of benzyl rings and modifying the chain length connecting the backbone to the aromatic moiety, we observe notable effects on fibril formation, molecular packing, and immunogenicity both in vitro and in vivo. Our results show that subtle chemical modifications are effective tools for designing tailored peptide nanomaterials with promising potential in vaccine delivery, tissue engineering, and regenerative medicine.

bioengineering↗

bioRxiv2024

Enantiomeric histidine-rich peptide coacervates enhance antigen delivery to T cells

Peptides and peptidomimetics that self-assemble via LLPS have recently emerged as building blocks for fabricating functional biomaterials due to their unique physicochemical properties and dynamic nature. One of lifes most distinctive signatures is its selectivity for chiral molecules and, to date, coacervates comprised of D-amino acids have not been reported. Here, we demonstrate that histidine-rich repeats of (GHGXY)4 (X=L/V/P) and their enantiomers undergo LLPS opening new avenues for enhancing coacervate stability. Through a series of biophysical studies, we find that LLPS kinetics, droplet size, fusion, and encapsulation efficiency are dictated by the primary sequence. Further, these coacervates can encapsulate therapeutic cargo which are then internalized via endocytic mechanisms. Finally, we show that the coacervates enhance antigen presentation to CD4+ and CD8+ T cells resulting in robust proliferation and production of functional cytokines. Collectively, our study describes the development and characterization of enantiomeric peptide coacervates as attractive vaccine delivery vehicles with tunable physicochemical properties. HIGHLIGHTSO_LID amino acid-peptides were used for the first time to construct phase separating coacervates C_LIO_LIChirality does not restrict LLPS or modulate other coacervate properties C_LIO_LIAntigen delivery using chiral coacervates enhances and prolongs presentation to T cells C_LI PROGRESS AND POTENTIALPeptides can undergo self-assembly via liquid-liquid phase separation (LLPS) to result in solute-rich coacervates that can serve as biomaterials. Using histidine-rich peptide repeats, this work demonstrates that peptides composed of entirely D-amino acids can form functional coaceravtes. The kinetics of LLPS and bulk properties of the droplets can be controlled through simple amino acid substitutions. The coacervates, while immunologically inert, exert an adjuvanting effect and enhance antigen presentation to T cells leading to proliferation and functional cytokine production. The materials showcased here possess high translational potential for combined delivery of immunomodulators and antigens for vaccine delivery against infectious diseases or cancer. The deliverables from this study will also inspire the development of chiral systems that will contribute to the knowledge of cellular processes associated with phase changes integral to both physiology and pathology.

bioengineering↗

bioRxiv2024

DAMP-inducing Peptide Nanofibers and PAMP Combination Adjuvants Boost Functional Lung Tissue-resident Memory CD4+ T Cell Responses

Vaccine adjuvants are typically composed of pathogen-associated molecular patterns (PAMPs) or danger-associated molecular patterns (DAMPs) that activate innate immune cells. Advances in basic immunology have demonstrated the need for various types of protective immunity, which are difficult to achieve with a single adjuvant. The FDA approval of multiple PAMP-DAMP combinations for clinical use has led to an increased momentum in the area in recent years. Here we report the use of DAMP-inducing peptide nanofibers (PNFs) and CL429 (PAMP) combinations as subunit boosters for Bacille Calmette-Guerin (BCG). We demonstrate that pulmonary boosting with PNFs and CL429 enhances the lung-resident memory phenotype, effector cytokine profiles, and transcription factor bias of antigen-specific CD4+T cell populations compared to PNFs alone. Importantly, the combination significantly improved the frequency of tissue-resident memory T (TRM) cells which, have been shown to provide superior protection compared to circulating memory T cells. Interestingly, the T helper (Th) subset profile was driven in part driven by the route of vaccination resulting in a Th17 bias via a mucosal route or a Th1 bias when delivered intravenously. We show that following pulmonary administration, lung-resident antigen presenting cells (APCs) efficiently internalize PNFs and upregulate important co-stimulatory markers that drive T cell priming and activation. Our findings suggest that heterologous booster vaccines composed of DAMP-inducing PNFs and PAMP combinations can engage innate and adaptive immunity for generating TRM cells that protect against TB and potentially other respiratory diseases.

bioengineering↗

bioRxiv2023

Resolving the nanoscale structure of β-sheet assemblies using single-molecule orientation-localization microscopy

Synthetic peptides that self-assemble into cross-{beta} fibrils have remarkable utility as engineered biomaterials due to their modularity and biocompatibility, but their structural and morphological similarity to amyloid species has been a long-standing concern for their translation. Further, their polymorphs are difficult to characterize using spectroscopic and imaging techniques that rely on ensemble averaging to achieve high resolution. Here, we utilize single-molecule orientation-localization microscopy (SMOLM) to characterize fibrils formed by the designed amphipathic enantiomers, KFE8L and KFE8D, and the pathological amyloid-beta peptide A{beta}42. SMOLM reveals that the orientations of Nile red, as it transiently binds to both KFE8 and A{beta}42, are consistent with a helical (bilayer) ribbon structure and convey the precise tilt of the fibrils inner and outer backbones. SMOLM also finds polymorphic branched and curved morphologies of KFE8 whose backbones exhibit much more heterogeneity than those of more typical straight fibrils. Thus, SMOLM is a powerful tool to interrogate the structural differences and polymorphism between engineered and pathological cross {beta}-rich fibrils.

biophysics↗

bioRxiv2021

Coiled Coil Crosslinked Alginate Hydrogels Dampen Macrophage-Driven Inflammation

Alginate hydrogels are widely used for tissue engineering and regenerative medicine due to their excellent biocompatibility. A facile and commonly used strategy to crosslink alginate is the addition of Ca2+ that leads to hydrogelation. However, extracellular Ca2+ is a secondary messenger in activating inflammasome pathways following physical injury or pathogenic insult leading to persistent inflammation and scaffold rejection. Here we present graft copolymers of charge complementary heterodimeric coiled coil (CC) peptides and alginate that undergo supramolecular self-assembly to form Ca2+ free alginate hydrogels. The formation of heterodimeric CCs was confirmed using circular dichroism spectroscopy and scanning electron microscopy revealed a significant difference in pore size between Ca2+ and CC crosslinked gels. The resulting hydrogels were self-supporting and display shear-thinning and shear-recovery properties. In response to lipopolysaccharide (LPS) stimulation, peritoneal macrophages and bone marrow derived dendritic cells cultured in the CC crosslinked gels exhibited a 10-fold reduction in secretion of the proinflammatory cytokine IL-1{beta} compared to Ca2+ crosslinked gels. A similar respose was also observed in vivo upon peritoneal delivery of Ca2+ or CC crosslinked gels. Analysis of peritoneal lavage showed that macrophages in mice injected with Ca2+ crosslinked gels display a more inflammatory phenotype compared to macrophages from mice injected with CC crosslinked gels. These results suggest that CC peptides by virtue of their tunable sequence-structure-function relationship and mild gelation conditions are promising alternative crosslinkers for alginate and other biopolymer scaffolds used in tissue engineering.

bioengineering↗