Pub Date : 2024-08-06DOI: 10.1101/2024.08.05.606187
Max Dongsheng Yin, Olivier N. Lemaire, José Guadalupe Rosas-Jiménez, Mélissa Belhamri, Anna Shevchenko, Gerhard Hummer, Tristan Wagner, Bonnie J. Murphy
In the ancient microbial Wood-Ljungdahl pathway, CO2 is fixed in a multi-step process with acetyl-CoA synthesis at the bifunctional carbon monoxide dehydrogenase/acetyl-CoA synthase complex (CODH/ACS). Here, we present catalytic snapshots of the CODH/ACS from the gas-converting acetogen Clostridium autoethanogenum, characterizing the molecular choreography of the overall reaction including electron transfer to the CODH for CO2 reduction, methyl transfer from the corrinoid iron-sulfur protein (CoFeSP) partner to the ACS active site and acetyl-CoA production. Unlike CODH, the multidomain ACS undergoes large conformational changes to form an internal connection to the CODH active site, accommodate the CoFeSP for methyl transfer and protect the reaction intermediates. Altogether, the structures allow us to draw a detailed reaction mechanism of this enzyme crucial for CO2 fixation in anaerobic organisms.
{"title":"Snapshots of acetyl-CoA synthesis, the final step of CO2 fixation in the Wood-Ljungdahl pathway","authors":"Max Dongsheng Yin, Olivier N. Lemaire, José Guadalupe Rosas-Jiménez, Mélissa Belhamri, Anna Shevchenko, Gerhard Hummer, Tristan Wagner, Bonnie J. Murphy","doi":"10.1101/2024.08.05.606187","DOIUrl":"https://doi.org/10.1101/2024.08.05.606187","url":null,"abstract":"In the ancient microbial Wood-Ljungdahl pathway, CO<sub>2</sub> is fixed in a multi-step process with acetyl-CoA synthesis at the bifunctional carbon monoxide dehydrogenase/acetyl-CoA synthase complex (CODH/ACS). Here, we present catalytic snapshots of the CODH/ACS from the gas-converting acetogen Clostridium autoethanogenum, characterizing the molecular choreography of the overall reaction including electron transfer to the CODH for CO<sub>2</sub> reduction, methyl transfer from the corrinoid iron-sulfur protein (CoFeSP) partner to the ACS active site and acetyl-CoA production. Unlike CODH, the multidomain ACS undergoes large conformational changes to form an internal connection to the CODH active site, accommodate the CoFeSP for methyl transfer and protect the reaction intermediates. Altogether, the structures allow us to draw a detailed reaction mechanism of this enzyme crucial for CO<sub>2</sub> fixation in anaerobic organisms.","PeriodicalId":501048,"journal":{"name":"bioRxiv - Biophysics","volume":"25 1","pages":""},"PeriodicalIF":0.0,"publicationDate":"2024-08-06","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"141945306","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2024-08-06DOI: 10.1101/2024.08.04.606536
Jonathan Remis, Petar N Petrov, Jessie T Zhang, Jeremy J Axelrod, Hang Cheng, Shahar Sandhaus, Holger Mueller, Robert M Glaeser
Apoferritin (apoF) is commonly used as a test specimen in single-particle electron cryo-microscopy (cryo-EM), since it consistently produces density maps that go to 0.3 nm resolution or higher. When we imaged apoF with a laser phase plate (LPP), however, we observed more severe particle-to-particle variation in the images than we had previously thought to exist. Similarly, we found that images of ribulose bisphosphate carboxylase/oxygenase (rubisco) also exhibited a much greater amount of heterogeneity than expected. By comparison to simulations of images, we verified that the heterogeneity is not explained by the known features of the LPP, shot noise, or differences in particle orientation. We also demonstrate that our specimens are comparable to those previously used in the literature, based on using the final-reconstruction resolution as the metric for evaluation. All of this leads us to the hypothesis that the heterogeneity is due to damage that has occurred either during purification of the specimen or during preparation of the grids. It is not, however, our goal to explain the causes of heterogeneity; rather, we report that using the LPP has made the apparent damage too obvious to be ignored. In hindsight, similar heterogeneity can be seen in images of apoF and the 20S proteasome which others had recorded with a Volta phase plate. We therefore conclude that the increased contrast of phase-plate images (at low spatial frequencies) should also make it possible to visualize, on a single-particle basis, various forms of biologically functional heterogeneity in structure that had previously gone unnoticed.
{"title":"Cryo-EM phase-plate images reveal unexpected levels of apparent specimen damage","authors":"Jonathan Remis, Petar N Petrov, Jessie T Zhang, Jeremy J Axelrod, Hang Cheng, Shahar Sandhaus, Holger Mueller, Robert M Glaeser","doi":"10.1101/2024.08.04.606536","DOIUrl":"https://doi.org/10.1101/2024.08.04.606536","url":null,"abstract":"Apoferritin (apoF) is commonly used as a test specimen in single-particle electron cryo-microscopy (cryo-EM), since it consistently produces density maps that go to 0.3 nm resolution or higher. When we imaged apoF with a laser phase plate (LPP), however, we observed more severe particle-to-particle variation in the images than we had previously thought to exist. Similarly, we found that images of ribulose bisphosphate carboxylase/oxygenase (rubisco) also exhibited a much greater amount of heterogeneity than expected. By comparison to simulations of images, we verified that the heterogeneity is not explained by the known features of the LPP, shot noise, or differences in particle orientation. We also demonstrate that our specimens are comparable to those previously used in the literature, based on using the final-reconstruction resolution as the metric for evaluation. All of this leads us to the hypothesis that the heterogeneity is due to damage that has occurred either during purification of the specimen or during preparation of the grids. It is not, however, our goal to explain the causes of heterogeneity; rather, we report that using the LPP has made the apparent damage too obvious to be ignored. In hindsight, similar heterogeneity can be seen in images of apoF and the 20S proteasome which others had recorded with a Volta phase plate. We therefore conclude that the increased contrast of phase-plate images (at low spatial frequencies) should also make it possible to visualize, on a single-particle basis, various forms of biologically functional heterogeneity in structure that had previously gone unnoticed.","PeriodicalId":501048,"journal":{"name":"bioRxiv - Biophysics","volume":"25 1","pages":""},"PeriodicalIF":0.0,"publicationDate":"2024-08-06","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"141945302","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2024-08-06DOI: 10.1101/2024.08.05.606641
Peter-Rory Hall, Thibault Jouen-Tachoire, Marcus Schewe, Peter Proks, Thomas Baukrowitz, Elisabeth P Carpenter, Simon Newstead, Karin Rodstrom, Stephen J Tucker
TASK-1 and TASK-3 are pH-sensitive Two-Pore Domain (K2P/KCNK) K+ channels. Their functional roles make them promising targets for the treatment of multiple disorders including sleep apnea, pain and atrial fibrillation. Rare genetic mutations in these channels are also associated with neurodevelopmental and hypertensive disorders. A recent crystal structure of TASK-1 revealed a lower 'X-gate' that is a hotspot for missense gain-of-function mutations associated with DDSA (Developmental Delay with Sleep Apnea). However, the structural basis for gating in TASK channels and how they sense extracellular pH to regulate gating have not been fully elucidated. Here, we resolve structures for both the human TASK-1 and TASK-3 channels by cryoEM, as well as for a recurrent TASK-3 variant (G236R) associated with KCNK9 Imprinting Syndrome (formerly referred to as Birk-Barel Syndrome). Combined with functional studies of the X-gating mechanism, these structures not only provide evidence for how a highly-conserved gating mechanism becomes defective in disease, but also provide further insight into the pathway of conformational changes that underlie the pH-dependent inhibition of TASK channel activity.
{"title":"Structures of TASK-1 and TASK-3 K2P channels provide insight into their gating and dysfunction in disease","authors":"Peter-Rory Hall, Thibault Jouen-Tachoire, Marcus Schewe, Peter Proks, Thomas Baukrowitz, Elisabeth P Carpenter, Simon Newstead, Karin Rodstrom, Stephen J Tucker","doi":"10.1101/2024.08.05.606641","DOIUrl":"https://doi.org/10.1101/2024.08.05.606641","url":null,"abstract":"TASK-1 and TASK-3 are pH-sensitive Two-Pore Domain (K2P/KCNK) K+ channels. Their functional roles make them promising targets for the treatment of multiple disorders including sleep apnea, pain and atrial fibrillation. Rare genetic mutations in these channels are also associated with neurodevelopmental and hypertensive disorders. A recent crystal structure of TASK-1 revealed a lower 'X-gate' that is a hotspot for missense gain-of-function mutations associated with DDSA (Developmental Delay with Sleep Apnea). However, the structural basis for gating in TASK channels and how they sense extracellular pH to regulate gating have not been fully elucidated. Here, we resolve structures for both the human TASK-1 and TASK-3 channels by cryoEM, as well as for a recurrent TASK-3 variant (G236R) associated with KCNK9 Imprinting Syndrome (formerly referred to as Birk-Barel Syndrome). Combined with functional studies of the X-gating mechanism, these structures not only provide evidence for how a highly-conserved gating mechanism becomes defective in disease, but also provide further insight into the pathway of conformational changes that underlie the pH-dependent inhibition of TASK channel activity.","PeriodicalId":501048,"journal":{"name":"bioRxiv - Biophysics","volume":"11 1","pages":""},"PeriodicalIF":0.0,"publicationDate":"2024-08-06","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"141945210","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2024-08-06DOI: 10.1101/2024.08.01.606085
Caelen G Boucher-Bergstedt, Mark A Jankauski, Erick Johnson
Buzz pollination involves the release of pollen from, primarily, poricidal anthers through vibrations generated by certain bee species. Despite previous experimental and numerical studies, the intricacies of pollen dynamics within vibrating anthers remain elusive due to the challenges in observing these small-scale, opaque systems. This research employs the discrete element method (DEM) to simulate the pollen expulsion process in vibrating anthers. By exploring various frequencies and displacement amplitudes, a correlation between the maximum jerk of anther walls and the initial rate of pollen expulsion is observed under translating oscillations. This study highlights that while increased vibration intensity enhances pollen release, the rate of increase diminishes at higher intensities. Our findings also reveal the significant role of pollen-pollen interactions, which account for upwards of one-third of the total collisions. Comparisons between poricidal and pseudoporicidal anther geometries suggest that pore size and shape also influence expulsion rates. This research provides a foundation for more comprehensive models that can incorporate additional factors such as cohesion, adhesion, and Coulomb forces, paving the way for deeper insights into the mechanics of buzz pollination and its variability across different anther types and vibration parameters.
{"title":"Buzz Pollination: Investigations of Pollen Expulsion using the Discrete Element Method","authors":"Caelen G Boucher-Bergstedt, Mark A Jankauski, Erick Johnson","doi":"10.1101/2024.08.01.606085","DOIUrl":"https://doi.org/10.1101/2024.08.01.606085","url":null,"abstract":"Buzz pollination involves the release of pollen from, primarily, poricidal anthers through vibrations generated by certain bee species. Despite previous experimental and numerical studies, the intricacies of pollen dynamics within vibrating anthers remain elusive due to the challenges in observing these small-scale, opaque systems. This research employs the discrete element method (DEM) to simulate the pollen expulsion process in vibrating anthers. By exploring various frequencies and displacement amplitudes, a correlation between the maximum jerk of anther walls and the initial rate of pollen expulsion is observed under translating oscillations. This study highlights that while increased vibration intensity enhances pollen release, the rate of increase diminishes at higher intensities. Our findings also reveal the significant role of pollen-pollen interactions, which account for upwards of one-third of the total collisions. Comparisons between poricidal and pseudoporicidal anther geometries suggest that pore size and shape also influence expulsion rates. This research provides a foundation for more comprehensive models that can incorporate additional factors such as cohesion, adhesion, and Coulomb forces, paving the way for deeper insights into the mechanics of buzz pollination and its variability across different anther types and vibration parameters.","PeriodicalId":501048,"journal":{"name":"bioRxiv - Biophysics","volume":"45 1","pages":""},"PeriodicalIF":0.0,"publicationDate":"2024-08-06","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"141945209","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2024-08-06DOI: 10.1101/2024.08.05.606600
Syed Yunus Ali, Ashok Prasad, Dibyendu Das
The timings of several cellular events like cell lysis, cell division, or pore formation in endosomes are regulated by the time taken for the relevant proteins to cross a threshold in number or concentration. Since protein synthesis is stochastic, the threshold crossing time is a first passage problem. The exact distributions of these first passage processes have been obtained recently for unregulated and auto-regulated genes. Many proteins are however regulated by post-transcriptional regulation, controlled by small non-coding RNAs (sRNAs). Certain mathematical models of gene expression with post-transcriptional sRNA regulation have been recently exactly mapped to models without sRNA regulation. Utilizing this mapping and the exact distributions, we calculate exact results on fluctuations (full distribution, all cumulants, and characteristic times) of protein threshold crossing times in the presence of sRNA regulation. We derive two interesting predictions from these exact results. We show that the size of the fluctuation of the threshold crossing times have a non-monotonic U-shaped behavior as a function of the rates of binding and unbinding of the sRNA-mRNA complex. Thus there are optimal parameters that minimize noise. Furthermore, the fluctuations in models with sRNA regulation may be higher or lower compared to the model without regulation, depending on the mean protein burst size.
{"title":"Exact distributions of threshold crossing times of proteins under post-transcriptional regulation by small RNAs","authors":"Syed Yunus Ali, Ashok Prasad, Dibyendu Das","doi":"10.1101/2024.08.05.606600","DOIUrl":"https://doi.org/10.1101/2024.08.05.606600","url":null,"abstract":"The timings of several cellular events like cell lysis, cell division, or pore formation in endosomes are regulated by the time taken for the relevant proteins to cross a threshold in number or concentration. Since protein synthesis is stochastic, the threshold crossing time is a first passage problem. The exact distributions of these first passage processes have been obtained recently for unregulated and auto-regulated genes. Many proteins are however regulated by post-transcriptional regulation, controlled by small non-coding RNAs (sRNAs). Certain mathematical models of gene expression with post-transcriptional sRNA regulation have been recently exactly mapped to models without sRNA regulation. Utilizing this mapping and the exact distributions, we calculate exact results on fluctuations (full distribution, all cumulants, and characteristic times) of protein threshold crossing times in the presence of sRNA regulation. We derive two interesting predictions from these exact results. We show that the size of the fluctuation of the threshold crossing times have a non-monotonic U-shaped behavior as a function of the rates of binding and unbinding of the sRNA-mRNA complex. Thus there are optimal parameters that minimize noise. Furthermore, the fluctuations in models with sRNA regulation may be higher or lower compared to the model without regulation, depending on the mean protein burst size.","PeriodicalId":501048,"journal":{"name":"bioRxiv - Biophysics","volume":"25 1","pages":""},"PeriodicalIF":0.0,"publicationDate":"2024-08-06","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"141945208","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2024-08-06DOI: 10.1101/2024.08.05.606726
Chunhong Long, Hongqiong Liang, Biao Wan
In eukaryotes, DNA achieves a highly compact structure primarily due to its winding around the histone cores. The nature wrapping of DNA around histone core form a 1.7 left-handed superhelical turns, contributing to negative supercoiling in chromatin. On the contrary, negative supercoils generated behind the polymerase during transcription may play a role in triggering nucleosome reassembly. To elucidate how supercoils influence the dynamics of wrapping of DNA around the histone cores, we developed a novel model to simulate the intricate interplay between DNA and histone. Our simulations revealed that both positively and negatively supercoiled DNAs are capable of wrapping around histone cores to adopt the nucleosome conformation. Most of all, our findings confirmed a preference for negative supercoiled DNA during nucleosome wrapping, and revealed that the both of the negative writhe and twist are comparatively beneficial to the formation of the DNA wrapping around histone. This advancement in the understanding of spontaneously nucleosome formation may provide insights into the intricate dynamics of chromatin assembly and its diverse functions. Our model thus can be further utilized to simulate the formations of multi-nucleosomes during re-assembling of the chromatin fiber, which will significantly enhance the understanding of the fundamental mechanisms governing the structure and function of chromatin.
在真核生物中，DNA 的结构非常紧凑，这主要是由于它缠绕在组蛋白核心上。DNA 围绕组蛋白核心的自然缠绕形成了 1.7 个左旋超螺旋转折，促成了染色质中的负超螺旋。相反，在转录过程中聚合酶后面产生的负超螺旋可能在触发核小体重新组装方面发挥作用。为了阐明超螺旋如何影响 DNA 围绕组蛋白核心的缠绕动态，我们开发了一个新模型来模拟 DNA 和组蛋白之间错综复杂的相互作用。我们的模拟结果表明，正向和负向超螺旋DNA都能缠绕组蛋白核心，从而形成核小体构象。最重要的是，我们的研究结果证实，在核小体包裹过程中，DNA更倾向于负超卷曲，并揭示了负蠕动和扭曲都相对有利于DNA包裹组蛋白的形成。对核小体自发形成的这一认识上的进步，可能有助于深入了解染色质组装的复杂动态及其各种功能。因此，我们的模型可进一步用于模拟染色质纤维重新组装过程中多核小体的形成，这将大大加深对染色质结构和功能基本机制的理解。
{"title":"DNA spontaneously wrapping around a histone core prefers negative supercoiling: A Brownian dynamics study","authors":"Chunhong Long, Hongqiong Liang, Biao Wan","doi":"10.1101/2024.08.05.606726","DOIUrl":"https://doi.org/10.1101/2024.08.05.606726","url":null,"abstract":"In eukaryotes, DNA achieves a highly compact structure primarily due to its winding around the histone cores. The nature wrapping of DNA around histone core form a 1.7 left-handed superhelical turns, contributing to negative supercoiling in chromatin. On the contrary, negative supercoils generated behind the polymerase during transcription may play a role in triggering nucleosome reassembly. To elucidate how supercoils influence the dynamics of wrapping of DNA around the histone cores, we developed a novel model to simulate the intricate interplay between DNA and histone. Our simulations revealed that both positively and negatively supercoiled DNAs are capable of wrapping around histone cores to adopt the nucleosome conformation. Most of all, our findings confirmed a preference for negative supercoiled DNA during nucleosome wrapping, and revealed that the both of the negative writhe and twist are comparatively beneficial to the formation of the DNA wrapping around histone. This advancement in the understanding of spontaneously nucleosome formation may provide insights into the intricate dynamics of chromatin assembly and its diverse functions. Our model thus can be further utilized to simulate the formations of multi-nucleosomes during re-assembling of the chromatin fiber, which will significantly enhance the understanding of the fundamental mechanisms governing the structure and function of chromatin.","PeriodicalId":501048,"journal":{"name":"bioRxiv - Biophysics","volume":"47 1","pages":""},"PeriodicalIF":0.0,"publicationDate":"2024-08-06","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"141945305","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2024-08-05DOI: 10.1101/2024.08.04.606526
Denys Biriukov, Zuzana Osifova, Man Nguyen Thi Hong, Philip E Mason, Martin Dracinsky, Pavel Jungwirth, Jan Heyda, Mattia I Morandi, Mario Vazdar
The phenomenon of like-charge pairing of hydrated ions is a physical manifestation of the unique solvation properties of certain ion pairs in water. Water's high dielectric constant and related ion screening capability significantly influence the interaction between like-charged ions, with the possibility to transform it - in some cases - from repulsion to attraction. Guanidinium cations (Gdm+) represent a quintessential example of such like-charge pairing due to their specific geometry and charge distribution. In this work, we present experimental quantification of Gdm+ - Gdm+ contact ion pairing in water utilizing nuclear magnetic resonance (NMR) spectroscopy experiments complemented by molecular dynamics (MD) simulations and density functional theory (DFT) calculations. The observed interaction is very weak - about -0.5 kJ mol-1 - which aligns with theoretical estimation from MD simulations. We also contrast the behavior of Gdm+ with NH4+ cations, which do no exhibit contact ion pairing in water. DFT calculations predict that the NMR chemical shift of Gdm+ dimers is smaller than that of monomers, in agreement with NMR titration curves that display a nonlinear Langmuir-like behavior. Additionally, we conducted cryo-electron microscopy experiments on oligoarginines R9, which (unlike nona-lysines K9) exhibit aggregation in water. This points again to like charge pairing of the guanidinium side chain groups, as corroborated also by molecular dynamics simulations of these peptides in water.
{"title":"The Origins of Arginine \"Magic\": Guanidinium Like-Charge Ion Pairing and Oligoarginine Aggregation in Water by NMR, Cryoelectron Microscopy, and Molecular Dynamics Simulations","authors":"Denys Biriukov, Zuzana Osifova, Man Nguyen Thi Hong, Philip E Mason, Martin Dracinsky, Pavel Jungwirth, Jan Heyda, Mattia I Morandi, Mario Vazdar","doi":"10.1101/2024.08.04.606526","DOIUrl":"https://doi.org/10.1101/2024.08.04.606526","url":null,"abstract":"The phenomenon of like-charge pairing of hydrated ions is a physical manifestation of the unique solvation properties of certain ion pairs in water. Water's high dielectric constant and related ion screening capability significantly influence the interaction between like-charged ions, with the possibility to transform it - in some cases - from\u0000repulsion to attraction. Guanidinium cations (Gdm+) represent a quintessential example of such like-charge pairing due to their specific geometry and charge distribution.\u0000In this work, we present experimental quantification of Gdm+ - Gdm+ contact ion pairing in water utilizing nuclear magnetic resonance (NMR) spectroscopy experiments complemented by molecular dynamics (MD) simulations and density functional theory (DFT) calculations. The observed interaction is very weak - about -0.5 kJ mol-1 - which aligns with theoretical estimation from MD simulations. We also contrast the behavior of Gdm+ with NH4+ cations, which do no exhibit contact ion pairing in water.\u0000DFT calculations predict that the NMR chemical shift of Gdm+ dimers is smaller than that of monomers, in agreement with NMR titration curves that display a nonlinear Langmuir-like behavior. Additionally, we conducted cryo-electron microscopy experiments on oligoarginines R9, which (unlike nona-lysines K9) exhibit aggregation in water. This points again to like charge pairing of the guanidinium side chain groups, as corroborated also by molecular dynamics simulations of these peptides in water.","PeriodicalId":501048,"journal":{"name":"bioRxiv - Biophysics","volume":"45 1","pages":""},"PeriodicalIF":0.0,"publicationDate":"2024-08-05","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"141945308","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2024-08-05DOI: 10.1101/2024.08.01.606202
Amaury Perez-Tirado, Ulla Unkelbach, Tabea A. Oswald, Johannes Rheinlaender, Tilman E. Schaeffer, Markus Mukenhirn, Alf Honigmann, Andreas Janshoff
Investigations of the response of curved epithelia derived from MDCK-II cells to external deformation involved indentation-relaxation experiments using colloidal probe microscopy. Notably, hemicysts exhibited lower tissue tension, greater compliance, and increased fluidity compared to cysts. The primary response to deformation turned out to be the in-plane expansion of the basal cortex/membrane of cells. Additionally, drug treatments applied to curved tissue, along with deformation of tailored mutants (such as E-cadherin knockout), revealed that tissue compliance over short time scales is influenced by an interplay of viscoelastic properties in individual cells, their apical-basal polarity, superelasticity of the shell, and excess interfacial area. Meanwhile, tissue resilience predominantly depends on the integrity of cell-cell contacts.
{"title":"Mechanical Implications of Cellular Viscoelasticity, Cortex Polarity, Superelasticity, and Cell-Cell Junctions in Curved Tissues","authors":"Amaury Perez-Tirado, Ulla Unkelbach, Tabea A. Oswald, Johannes Rheinlaender, Tilman E. Schaeffer, Markus Mukenhirn, Alf Honigmann, Andreas Janshoff","doi":"10.1101/2024.08.01.606202","DOIUrl":"https://doi.org/10.1101/2024.08.01.606202","url":null,"abstract":"Investigations of the response of curved epithelia derived from MDCK-II cells to external deformation involved indentation-relaxation experiments using colloidal probe microscopy. Notably, hemicysts exhibited lower tissue tension, greater compliance, and increased fluidity compared to cysts. The primary response to deformation turned out to be the in-plane expansion of the basal cortex/membrane of cells. Additionally, drug treatments applied to curved tissue, along with deformation of tailored mutants (such as E-cadherin knockout), revealed that tissue compliance over short time scales is influenced by an interplay of viscoelastic properties in individual cells, their apical-basal polarity, superelasticity of the shell, and excess interfacial area. Meanwhile, tissue resilience predominantly depends on the integrity of cell-cell contacts.","PeriodicalId":501048,"journal":{"name":"bioRxiv - Biophysics","volume":"102 1","pages":""},"PeriodicalIF":0.0,"publicationDate":"2024-08-05","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"141945307","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2024-08-05DOI: 10.1101/2024.08.04.606547
Chenlin Lu, Malcolm L. Wells, Andrew Reckers, Anum Glasgow
While bioinformatics reveals patterns in protein sequences and structural biology methods elucidate atomic details of protein structures, it is difficult to attain equally high-resolution energetic information about protein conformational ensembles. We present PIGEON-FEATHER, a method for calculating free energies of opening (∆Gop) at single- or near-single-amino acid resolution for protein ensembles of all sizes from hydrogen exchange/mass spectrometry (HX/MS) data. PIGEON-FEATHER disambiguates and reconstructs all experimentally measured isotopic mass envelopes using a Bayesian Monte Carlo sampling approach. We applied PIGEON-FEATHER to reveal how E. coli and human dihydrofolate reductase orthologs (ecDHFR, hDHFR) have evolved distinct ensembles tuned to their catalytic cycles, and how two competitive inhibitors of ecDHFR arrest its ensemble in different ways. Extending the method to a large protein-DNA complex, we mapped ligand-induced ensemble reweighting in the E. coli lac repressor to understand the functional switching mechanism crucial for transcriptional regulation.
{"title":"Site-resolved energetic information from HX/MS experiments","authors":"Chenlin Lu, Malcolm L. Wells, Andrew Reckers, Anum Glasgow","doi":"10.1101/2024.08.04.606547","DOIUrl":"https://doi.org/10.1101/2024.08.04.606547","url":null,"abstract":"While bioinformatics reveals patterns in protein sequences and structural biology methods elucidate atomic details of protein structures, it is difficult to attain equally high-resolution energetic information about protein conformational ensembles. We present PIGEON-FEATHER, a method for calculating free energies of opening (∆Gop) at single- or near-single-amino acid resolution for protein ensembles of all sizes from hydrogen exchange/mass spectrometry (HX/MS) data. PIGEON-FEATHER disambiguates and reconstructs all experimentally measured isotopic mass envelopes using a Bayesian Monte Carlo sampling approach. We applied PIGEON-FEATHER to reveal how E. coli and human dihydrofolate reductase orthologs (ecDHFR, hDHFR) have evolved distinct ensembles tuned to their catalytic cycles, and how two competitive inhibitors of ecDHFR arrest its ensemble in different ways. Extending the method to a large protein-DNA complex, we mapped ligand-induced ensemble reweighting in the E. coli lac repressor to understand the functional switching mechanism crucial for transcriptional regulation.","PeriodicalId":501048,"journal":{"name":"bioRxiv - Biophysics","volume":"84 1","pages":""},"PeriodicalIF":0.0,"publicationDate":"2024-08-05","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"141945309","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2024-08-05DOI: 10.1101/2024.08.05.606589
Yohalie Kalukula, Marine Luciano, Guillaume Charras, David Brueckner, Sylvain Gabriele
Cell migration in narrow microenvironments is a hallmark of numerous physiological processes, involving successive cycles of confinement and release that drive significant morphological changes. However, it remains unclear whether migrating cells can retain a memory of their past morphological states, which could potentially enhance their navigation through confined spaces. By combining cell migration assays on standardized microsystems with biophysical modeling and biochemical perturbations, we demonstrate that local geometry governs these morphological switches, thereby facilitating cell passage through long and narrow gaps. We uncovered a long-term memory of past confinement events in migrating cells, with morphological states correlated across transitions through actin cortex remodeling. These findings suggest that mechanical memory in migrating cells plays an active role in their migratory potential in confined environments.
{"title":"The actin cortex acts as a mechanical memory of morphology in confined migrating cells","authors":"Yohalie Kalukula, Marine Luciano, Guillaume Charras, David Brueckner, Sylvain Gabriele","doi":"10.1101/2024.08.05.606589","DOIUrl":"https://doi.org/10.1101/2024.08.05.606589","url":null,"abstract":"Cell migration in narrow microenvironments is a hallmark of numerous physiological processes, involving successive cycles of confinement and release that drive significant morphological changes. However, it remains unclear whether migrating cells can retain a memory of their past morphological states, which could potentially enhance their navigation through confined spaces. By combining cell migration assays on standardized microsystems with biophysical modeling and biochemical perturbations, we demonstrate that local geometry governs these morphological switches, thereby facilitating cell passage through long and narrow gaps. We uncovered a long-term memory of past confinement events in migrating cells, with morphological states correlated across transitions through actin cortex remodeling. These findings suggest that mechanical memory in migrating cells plays an active role in their migratory potential in confined environments.","PeriodicalId":501048,"journal":{"name":"bioRxiv - Biophysics","volume":"58 1","pages":""},"PeriodicalIF":0.0,"publicationDate":"2024-08-05","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"141945310","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
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