Publications by Author: Celeste Kennard

Biotechnological innovations have vastly improved the capacity to perform large-scale protein studies, while the methods we have for identifying and quantifying individual proteins are still inadequate to perform protein sequencing at the single-molecule level. Nanopore-inspired systems devoted to understanding how single molecules behave have been extensively developed for applications in genome sequencing. These nanopore systems are emerging as prominent tools for protein identification, detection, and analysis, suggesting realistic prospects for novel protein sequencing. This review summarizes recent advances in biological nanopore sensors toward protein sequencing, from the identification of individual amino acids to the controlled translocation of peptides and proteins, with attention focused on device and algorithm development and the delineation of molecular mechanisms with the aid of simulations. Specifically, the review aims to offer recommendations for the advancement of nanopore-based protein sequencing from an engineering perspective, highlighting the need for collaborative efforts across multiple disciplines. These efforts should include chemical conjugation, protein engineering, molecular simulation, machine-learning-assisted identification, and electronic device fabrication to enable practical implementation in real-world scenarios.

G-protein-coupled receptors (GPCRs) mediate many critical physiological processes. Their spatial organization in plasma membrane (PM) domains is believed to encode signaling specificity and efficiency. However, the existence of domains and, crucially, the mechanism of formation of such putative domains remain elusive. Here, live-cell imaging (corrected for topography-induced imaging artifacts) conclusively established the existence of PM domains for GPCRs. Paradoxically, energetic coupling to extremely shallow PM curvature (<1 µm-1) emerged as the dominant, necessary and sufficient molecular mechanism of GPCR spatiotemporal organization. Experiments with different GPCRs, H-Ras, Piezo1 and epidermal growth factor receptor, suggest that the mechanism is general, yet protein specific, and can be regulated by ligands. These findings delineate a new spatiomechanical molecular mechanism that can transduce to domain-based signaling any mechanical or chemical stimulus that affects the morphology of the PM and suggest innovative therapeutic strategies targeting cellular shape.