Biology subjects

Polasa, A.

Publications and source records attributed to Polasa, A..

9 recordsLinked to original sources

bioRxiv2025

Benchmarking antigen-aware inverse folding methods for antibody design.

Computational antibody design has seen many recent advances pioneered via the use of language models and advanced structure prediction tools. Developing a de novo antibody against a specific antigen requires structural awareness that most language models lack. A prominent class of machine learning methods combining the best of language model and structural worlds is inverse folding. This approach aims to predict a sequence that would fit a given structure. Such methods are now increasingly used to predict alternate sequences given a structure of a binder. It is known that, just like language models, such methods have certain predictive power in identifying binders. Here we performed a set of tests to reveal where, if at all, such methods provide value in the realistic setting of antibody discovery.

bioinformatics↗

bioRxiv2025

nanoFOLD : sequence design of nanobodies via inverse folding

Antibodies devoid of light chains are a promising class of biotherapeutics. Computational methods that address these molecules are crucially needed to accelerate the traditional, long and expensive experimental process of their discovery. Inverse folding, wherein one is tasked to predict a sequence given molecular coordinates, is an established method in scaffold-based protein design. Here we develop an inverse folding method speci[fi]c to nanobodies. We demonstrate its application in nanobody-engineering scenarios of enriching binders from next-generation sequencing experiments and novel binder design.

bioinformatics↗

bioRxiv2024

Cholesterol Dependence on the Conformational Changes of Metabotropic Glutamate Receptor 1

Metabotropic glutamate receptors (mGluRs) are class C G protein-coupled receptors that function as obligate dimers in regulating neurotransmission and synaptic plasticity in the central nervous system. The mGluR1 subtype has been shown to be modulated by the membrane lipid environment, particularly cholesterol, though the molecular mechanisms remain elusive. In this study, we employed all-atom molecular dynamics simulations to investigate the effects of cholesterol on the conformational dynamics of the mGluR1 seven-transmembrane (7TM) domain in an inactive state model. Simulations were performed with three different cholesterol concentrations (0%, 10%, and 25%) in a palmitoyl-oleoyl phosphatidylcholine (POPC) lipid bilayer system. Our results demonstrate that cholesterol induces conformational changes in the mGluR1 dimer more significantly than in the individual protomers. Notably, cholesterol modulates the dynamics and conformations of the TM1 and TM2 helices at the dimer interface. Interestingly, an intermediate cholesterol concentration of 10% elicits more pronounced conformational changes compared to both cholesterol-depleted (0%) and cholesterol-enriched (25%) systems. Specific electrostatic interaction unique to the 10% cholesterol system further corroborate these conformational differences. Given the high sequence conservation of the 7TM domains across mGluR subtypes, the cholesterol-dependent effects observed in mGluR1 are likely applicable to other members of this receptor family. Our findings provide atomistic insights into how cholesterol modulates the conformational landscape of mGluRs, which could impact their function and signaling mechanisms.

biophysics↗

bioRxiv2024

Differential Behavior of Conformational Dynamics in Active and Inactive States of Cannabinoid Receptor 1 Revealed by Microsecond-Level Molecular Dynamics Simulation

Cannabinoid receptor 1 (CB1) is a G protein-coupled receptor (GPCR) that regulates critical physiological processes including pain, appetite, and cognition. Understanding the confor- mational dynamics of CB1 associated with transitions between inactive and active signaling states is imperative for developing targeted modulators. Using microsecond-level all-atom molecular dynamics (MD) simulations, we identified marked differences in the conformational ensembles of inactive and active CB1 in apo. The inactive state exhibited substantially in- creased structural heterogeneity and plasticity compared to the more rigidified active state in the absence of stabilizing ligands. Transmembrane helices TM3 and TM7 were identified as distinguishing factors modulating the state-dependent dynamics. TM7 displayed amplified fluctuations selectively in the inactive state simulations attributed to disruption of conserved electrostatic contacts anchoring it to surrounding helices in the active state. Additionally, we identified significant reorganizations in key salt bridge and hydrogen bond networks con- tributing to the CB1 activation/inactivation. For instance, D213-Y224 hydrogen bond and D184-K192 salt bridge showed marked rearrangements between the states. Collectively, these findings reveal the specialized role of TM7 in directing state-dependent CB1 dynamics through electrostatic switch mechanisms. By elucidating the intrinsic enhanced flexibility of inactive CB1, this study provides valuable insights into the conformational landscape enabling functional transitions. Our perspective advances understanding of CB1 activation mechanisms and offers opportunities for structure-based drug discovery targeting the state- specific conformational dynamics of this receptor. Graphic for manuscript O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=174 SRC="FIGDIR/small/589939v2_ufig1.gif" ALT="Figure 1"> View larger version (60K): org.highwire.dtl.DTLVardef@ade7d7org.highwire.dtl.DTLVardef@1500e0corg.highwire.dtl.DTLVardef@ca4b04org.highwire.dtl.DTLVardef@10317eb_HPS_FORMAT_FIGEXP M_FIG For Table of Contents Only C_FIG

biophysics↗

bioRxiv2022

Deciphering the Inter-domain Decoupling in the Gram-negative Bacterial Membrane Insertase

YidC is a membrane protein that plays an important role in inserting newly generated proteins into lipid membranes. The Sec-dependent complex is responsible for inserting proteins into the lipid bilayer, and this process is facilitated by YidC in bacteria. In addition, YidC acts as a chaperone during the folding process of proteins. Multiple investigations have conclusively shown that the gram-positive bacterial YidC has Sec-independent insertion mechanisms. Through the use of microsecond-level allatom molecular dynamics (MD) simulations, we have carried out the first in-depth investigation of the YidC protein originating from gram-negative bacteria. This research sheds light on the significance of multiple domains of YidC structure at an atomic level by utilizing equilibrium MD simulations. Specifically, in this research, multiple models of YidC embedded in the lipid bilayer were constructed to characterize the critical role of the C2 loop and the periplasmic domain present in gram-negative YidC, which is absent in its gram-positive counterpart. Based on our results, the C2 loop is responsible for the overall stabilization of the protein, most notably in the transmembrane region, and it also has an allosteric influence on the periplasmic domain. We have found critical inter- and intra-domain interactions that contribute to the stability of the protein and its function. Finally, our study provides a hypothetical Sec-independent insertion mechanism for gram-negative bacterial YidC.

biophysics↗

bioRxiv2022

An Investigation of the YidC-Mediated Membrane Insertion of Pf3 Coat Protein Using Molecular Dynamics Simulations

YidC is a membrane protein that facilitates the insertion of newly synthesized proteins into lipid membranes. Through YidC, proteins are inserted into the lipid bilayer via the SecYEG-dependent complex. Additionally, YidC functions as a chaperone in protein folding processes. Several studies have provided evidence of its independent insertion mechanism. However, the mechanistic details of the YidC independent protein insertion mechanism remain elusive at the molecular level. This study elucidates the insertion mechanism of YidC at an atomic level through a combination of equilibrium and non-equilibrium molecular dynamics (MD) simulations. Different docking models of YidC-Pf3 in the lipid bilayer were built in this study to better understand the insertion mechanism. To conduct a complete investigation of the conformational difference between the two docking models developed, we used classical molecular dynamics simulations supplemented with a non-equilibrium technique. Our findings indicate that the YidC transmembrane (TM) groove is essential for this high-affinity interaction and that the hydrophilic nature of the YidC groove plays an important role in protein transport across the cytoplasmic membrane bilayer to the periplasmic side. At different stages of the insertion process, conformational changes in YidCs TM domain and membrane core have a mechanistic effect on the Pf3 coat. Furthermore, during the insertion phase, the hydration and dehydration of the YidCs hydrophilic groove are critical. These demonstrate that Pf3 interactions with the membrane and YidC vary in different conformational states during the insertion process. Finally, this extensive study directly confirms that YidC functions as an independent insertase.

biophysics↗

bioRxiv2021

Computational Elucidation of Recombinant Fusion Protein Effect on Peptide-Directed Nanoparticles

Nanoparticles synthesized using various peptides have optimized properties and functional abilities which can be achieved via peptide flexibility and site specificity. Using peptide Pd4 and other alanine substitution combinations of Pd4 attached to a green fluorescent protein (GFPuv), nanoparticles with well-defined sizes that are soluble in aqueous solutions can be produced. In this study, extensive molecular dynamics simulations explored the structural and functional differences between the free peptides and the peptides bound to the GFPuv used in nanoparticle production. Binding affinities of histidines of Pd4 peptide and its two mutants A6 and A11 to a palladium atom were calculated using the free energy perturbation method. Interestingly, the average particle sizes obtained from transmission electron microscopy (TEM) images correlated with our calculated free energies of different peptide sequences. Remarkably, when the peptide was bound to GFPuv, the free energies of histidine were very similar in the wild-type and other mutated peptides. However, this trend is not observed with free peptide simulations, where binding affinities differ by mutation of histidine residues. This study describes, at a molecular level, the role of amino acid sequence on binding affinity of the peptide to the surface of the palladium particles, and the functional ability of the GFPuv protein controlling these free energies irrespective of peptide sequence. Our study will provide a framework for designing free and protein attached peptides that facilitate peptide-mediated nanoparticle formation with well-regulated properties.

bioengineering↗

bioRxiv2020

Differential Dynamic Behavior of Prefusion Spike Proteins of SARS Coronaviruses 1 and 2

The coronavirus spike protein, which binds to the same human receptor in both SARS-CoV-1 and 2, has been implied to be a potential source of their differential transmissibility. However, the mechanistic details of spike protein binding to its human receptor remain elusive at the molecular level. Here, we have used an extensive set of unbiased and biased microsecond-level all-atom molecular dynamics (MD) simulations of SARS-CoV-1 and 2 spike proteins to determine the differential dynamic behavior of prefusion spike protein structure in the two viruses. Our results indicate that the active form of the SARS-CoV-2 spike protein is more stable than that of SARS-CoV-1 and the energy barrier associated with the activation is higher in SARS-CoV-2. Our results also suggest that not only the receptor binding domain (RBD) but also other domains such as the N-terminal domain (NTD) could play a role in the differential binding behavior of SARS-CoV-1 and 2 spike proteins.

biophysics↗

bioRxiv2019

Elucidating the Molecular Basis of pH Activation of an Engineered Mechanosensitive Channel

Mechanosensitive (MS) channels detect and respond to changes in the pressure profile of cellular membranes and transduce the mechanical energy into electrical and/or chemical signals. However, by re-engineering the MS channels, chemical signals such as pH change can trigger the activation of some MS channels. This paper elucidate the activation mechanism of an engineered MS channel of large conductance (MscL) at an atomic level through a combination of equilibrium, non-equilibrium, biased, and unbiased molecular dynamics (MD) simulations for the first time. Comparing the wild-type and engineered MscL activation processes at an atomic level suggests that the two systems are likely to be associated with different active states and different transition pathways. These findings indicate that (1) periplasmic loops play a key role in the activation process of MscL, (2) the loss of various hydrogen bonds and salt bridge interactions in the engineered MscL channel causes the spontaneous opening of the channel, and (3) the most significant interactions lost during the activation process are those between the transmembrane (TM) helices 1 and 2 (TM1 and TM2) in engineered MscL channel. In this research, the orientation-based biasing approach for producing and optimizing an open MscL model is a promising way to characterize unknown protein functional states and to research the activation processes in ion channels. String method with swarms of trajectories (SMwST) was used to identify the optimal transition pathway and elucidate the activation mechanism of the engineered MscL. Finally, the free energy profile of engineered MscL associated with the activation process using a novel along-the-path free energy calculation approach is constructed. This work paves the way for a computational framework for the studies aimed at designing pH-triggered channel-functionalized drug delivery liposomes.

biophysics↗