Biology subjects

Popa, I.

Publications and source records attributed to Popa, I..

4 recordsLinked to original sources

bioRxiv2026

Precise Alternation Between Image-Forming Sample Planes Enables Quantitative Monitoring of Receptor-Arrestin Interaction Dynamics at the Plasma Membrane of Live Cells

Investigations of G protein-coupled receptors (GPCRs) interactions with non-visual arrestins in living cells are essential to understanding the complex molecular mechanisms of GPCR-based signaling. Quantitative analysis of these interactions remains challenging in live cells, particularly when attempting to repeatedly image distinct cellular regions with high precision. Here, we describe the implementation of an optical imaging stabilization approach that integrates the recently developed Focal Readjustment for Enhanced Vertical Resolution (FREVR) technology into a multiphoton microscope, enabling high-precision alternation between image-forming sample planes with < 20-nanometer repeatability and stability over time. Using this setup, we monitored the dynamic recruitment of arrestin-2 (Arr2) to the plasma membrane of HEK-293 cells expressing muscarinic acetylcholine M2 receptors (M2R) by alternatively imaging distinct planes of interest, the basolateral membrane and a membrane cross-section. Following stimulation of M2R by agonist ligand, we observed a pronounced redistribution of cytoplasmic arrestin-2 toward the plasma membrane in both cellular cross-sections and at the basolateral membrane. This method enables direct comparison of receptor and arrestin dynamics across regions of individual cells with very high precision, eliminating the need for averaging over numerous cells in order to denoise biologically relevant signals, and thereby capturing physiological cell-to-cell variability.

biophysics↗

bioRxiv2019

Mechanical reinforcement of protein L from Finegoldia magna points to a new bind-and-search mechanism

Several significant bacterial pathogens in humans secrete surface proteins that bind antibodies in order to protect themselves from the adaptive immune response and have evolved to operate under the mechanical sheer generated by mucus flow, coughing or urination. Protein L is secreted by Finegoldia magna and has several antibody-binding domains. These domains have two antibody-binding sites with vastly different avidity and the function of the second weaker binding interface is currently unknown. Here we use magnetic tweezers and covalent attachment via HaloTag and SpyTag to expose Protein L to unfolding forces in the absence and presence of antibody-ligands. We find that antibody binding increases the mechanical stability of protein L. Using the change in mechanical stability as a binding reporter, we determined that the low-avidity binding site is acting as a mechano-sensor. We propose a novel mechanism where the high-avidity binding site engages the tether, while the low-avidity binding site acts as a mechano-sensor, allowing bacteria to sample the antibody surface concentration and localize its search during successful binding under strain.\n\nSignificanceIt is well known that bacteria have an arsenal of tools to invade and to avoid dislocation. Based on the molecular response of a protein used by anaerobic bacteria to attach to antibodies and disrupt the immune system, here we report on a force-sensor-like behavior, triggered by antibody clusters and force. This pseudo-catch bond between bacteria and antibodies is activated through a second binding site which has lower avidity to antibodies, and which acts as a mechanical sensor, potentially regulating the search radii of the bacterium. Understanding of the bacteria attachment mechanism is of great importance toward developing new antibiotics and mechano-active drugs.

biophysics↗

bioRxiv2019

Chemical unfolding of protein domains induces shape change in programmed protein hydrogels

Programmable behavior combined with tailored stiffness and tunable biomechanical response are key requirements for developing successful materials. However, these properties are still an elusive goal for protein-based biomaterials. Here, we present a new method based on protein-polymer interactions, to manipulate the stiffness of protein-based hydrogels made from bovine serum albumin (BSA) by using polyelectrolytes such as poly(Ethelene)imine (PEI) and poly-L-lysine (PLL) at various concentrations. This approach confers protein-hydrogels tunable wide-range stiffness, from ~ 10 - 60 kPa when treated with PEI, without affecting the protein mechanics and nanostructure. We ascribe the increase in stiffness to the synergistic effect of the non-covalent electrostatic polymer-protein interaction, as well as the polymer-shell that stabilizes the protein domains nanomechanics. We use the 6-fold increase in stiffness induced by PEI to program BSA-hydrogels in various shapes. By utilizing the characteristic protein unfolding we can induce reversible shape-memory behavior of these composite materials using chemical denaturing solutions. We anticipate this novel approach based on protein engineering and polymer reinforcing will enable the development and investigation of new smart biomaterials and extend protein hydrogel capabilities beyond their conventional applications.

biophysics↗

bioRxiv2018

Modeling Protein-based Hydrogels under Force

Hydrogels made from structured polyprotein domains combine the properties of cross-linked polymers with the unfolding phase transition. The use of protein hydrogels as an ensemble approach to study the physics of domain unfolding is limited by the lack of scaling tools and by the complexity of the system. Here we propose a model to describe the biomechanical response of protein hydrogels based on the unfolding and extension of protein domains under force. Our model takes into account the contributions on the network dynamics of the molecules inside the gels, which have random cross-linking points and random topology. This model reproduces reported macroscopic visco-elastic effects and constitutes an important step toward using rheometry on protein hydrogels to scale down to the average mechanical response of protein molecules.

biophysics↗