Pub Date : 2024-09-10DOI: 10.1101/2024.09.09.612055
Dillon Balthrop, Deepesh Sigdel, Chunfeng Mao, Yuk-Ching Tse-Dinh, Maria Mills
Type IA topoisomerases relieve torsional stress in DNA by a strand-passage mechanism, using the strain in the DNA to drive relaxation. The topoisomerase IAs of the Mycobacterium genus have distinct C-terminal domains which are crucial for successful strand-passage. We used single-molecule magnetic tweezers to observe supercoil relaxation by wild type Mycobacterium smegmatis topoisomerase IA and two C-terminal truncation mutants. We recorded distinct behaviors from each truncation mutant. We calculated the free energy stored in the DNA as it is twisted under force to examine the differences between the proteins. Based on our results, we propose a modified model of the strand-passage cycle.
IA 型拓扑异构酶通过链传递机制缓解 DNA 中的扭转应力，利用 DNA 中的应变驱动松弛。分枝杆菌属的IA型拓扑异构酶具有不同的C端结构域，这些结构域对成功的链通过至关重要。我们使用单分子磁镊观察了野生型分枝杆菌拓扑异构酶IA和两个C端截断突变体的超螺旋松弛。我们记录了每个截断突变体的不同行为。我们计算了 DNA 受力扭曲时储存的自由能，以研究蛋白质之间的差异。基于我们的研究结果，我们提出了一个修改过的链-通道循环模型。
{"title":"Mycobacterial Topoisomerase I Energetically Suffers From C-Terminal Deletions","authors":"Dillon Balthrop, Deepesh Sigdel, Chunfeng Mao, Yuk-Ching Tse-Dinh, Maria Mills","doi":"10.1101/2024.09.09.612055","DOIUrl":"https://doi.org/10.1101/2024.09.09.612055","url":null,"abstract":"Type IA topoisomerases relieve torsional stress in DNA by a strand-passage mechanism, using the strain in the DNA to drive relaxation. The topoisomerase IAs of the Mycobacterium genus have distinct C-terminal domains which are crucial for successful strand-passage. We used single-molecule magnetic tweezers to observe supercoil relaxation by wild type <em>Mycobacterium smegmatis</em> topoisomerase IA and two C-terminal truncation mutants. We recorded distinct behaviors from each truncation mutant. We calculated the free energy stored in the DNA as it is twisted under force to examine the differences between the proteins. Based on our results, we propose a modified model of the strand-passage cycle.","PeriodicalId":501048,"journal":{"name":"bioRxiv - Biophysics","volume":"42 1","pages":""},"PeriodicalIF":0.0,"publicationDate":"2024-09-10","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142178247","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 : 2024-09-10DOI: 10.1101/2024.09.09.612012
Lingfeng Xue, Zigang Song, Qi Ouyang, Chen Song
Terahertz (THz) electromagnetic fields are increasingly recognized for their crucial roles in various aspects of medical research and treatment. Recent computational studies have demonstrated that THz waves can modulate ion channel function by interacting with either the channel proteins or the bound ions through distinct mechanisms. Here we outline a universal simulation protocol to identify the THz frequencies that may affect ion channels, which consists of frequency spectrum analysis and ion conductance analysis. Following this protocol, we studied the effect of THz field on a Cav channel and found a broad frequency band in 1 to 20 THz range. We believe that this protocol, along with the identified characteristic frequencies, will provide a theoretical foundation for future terahertz experimental studies.
{"title":"Protocol for Simulating the Effect of THz Electromagnetic Field on Ion Channels","authors":"Lingfeng Xue, Zigang Song, Qi Ouyang, Chen Song","doi":"10.1101/2024.09.09.612012","DOIUrl":"https://doi.org/10.1101/2024.09.09.612012","url":null,"abstract":"Terahertz (THz) electromagnetic fields are increasingly recognized for their crucial roles in various aspects of medical research and treatment. Recent computational studies have demonstrated that THz waves can modulate ion channel function by interacting with either the channel proteins or the bound ions through distinct mechanisms. Here we outline a universal simulation protocol to identify the THz frequencies that may affect ion channels, which consists of frequency spectrum analysis and ion conductance analysis. Following this protocol, we studied the effect of THz field on a Cav channel and found a broad frequency band in 1 to 20 THz range. We believe that this protocol, along with the identified characteristic frequencies, will provide a theoretical foundation for future terahertz experimental studies.","PeriodicalId":501048,"journal":{"name":"bioRxiv - Biophysics","volume":"1 1","pages":""},"PeriodicalIF":0.0,"publicationDate":"2024-09-10","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142178241","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 : 2024-09-10DOI: 10.1101/2024.09.08.611919
Miriam Rose Hood, Susan Marqusee
A protein′s energy landscape, all the accessible conformations, their populations, and their dynamics of interconversion, is encoded in its primary sequence. While we have a good understanding of how a protein′s primary sequence encodes its native state, we have a much weaker understanding of how sequence encodes the kinetic barriers such as unfolding and refolding. Here we have looked at two subtiliase homologs from the Bacillus subtilis, Intracellular Subtilisin Protease 1 (ISP1) and Subtilisin E (SbtE) that are expected to have very different dynamics. As an intracellular protein, ISP1 has a small pro-domain thought to act simply as a zymogen, whereas the extracellular SbtE has a large pro-domain required for folding. We examined the global and local energetics of the mature proteases and how each pro-domain impacts their landscapes. We find that ISP1′s pro-domain has limited impact on the energy landscape while the mature SbtE is thermodynamically unstable and kinetically trapped. The impact of the pro-domain has opposite effects on the flexibility of the core of the protein. ISP1′s core becomes more flexible while SbtE′s core becomes more rigid. ISP1 contains a conserved amino-acid insertion not present in extracellular subtilisin proteases, which points to a potential source for these differences. These homologs are an extreme example of how changes in the primary sequence can dramatically alter a proteins energy landscape, both stability and dynamics, and highlight the need for large scale, high throughput studies on the relationship between primary sequence and conformational dynamics.
{"title":"Exploring the sequence and structural determinants of the energy landscape from thermodynamically stable and kinetically trapped subtilisins: ISP1 and SbtE","authors":"Miriam Rose Hood, Susan Marqusee","doi":"10.1101/2024.09.08.611919","DOIUrl":"https://doi.org/10.1101/2024.09.08.611919","url":null,"abstract":"A protein′s energy landscape, all the accessible conformations, their populations, and their dynamics of interconversion, is encoded in its primary sequence. While we have a good understanding of how a protein′s primary sequence encodes its native state, we have a much weaker understanding of how sequence encodes the kinetic barriers such as unfolding and refolding. Here we have looked at two subtiliase homologs from the <em>Bacillus subtilis</em>, Intracellular Subtilisin Protease 1 (ISP1) and Subtilisin E (SbtE) that are expected to have very different dynamics. As an intracellular protein, ISP1 has a small pro-domain thought to act simply as a zymogen, whereas the extracellular SbtE has a large pro-domain required for folding. We examined the global and local energetics of the mature proteases and how each pro-domain impacts their landscapes. We find that ISP1′s pro-domain has limited impact on the energy landscape while the mature SbtE is thermodynamically unstable and kinetically trapped. The impact of the pro-domain has opposite effects on the flexibility of the core of the protein. ISP1′s core becomes more flexible while SbtE′s core becomes more rigid. ISP1 contains a conserved amino-acid insertion not present in extracellular subtilisin proteases, which points to a potential source for these differences. These homologs are an extreme example of how changes in the primary sequence can dramatically alter a proteins energy landscape, both stability and dynamics, and highlight the need for large scale, high throughput studies on the relationship between primary sequence and conformational dynamics.","PeriodicalId":501048,"journal":{"name":"bioRxiv - Biophysics","volume":"25 1","pages":""},"PeriodicalIF":0.0,"publicationDate":"2024-09-10","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142178208","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 : 2024-09-10DOI: 10.1101/2024.09.09.612134
Liam Haas-Neill, Deniz Meneksedag-Erol, Ayesha Chaudhry, Masha Novoselova, Qirat F. Ashraf, Elvin D. de Araujo, Derek J. Wilson, Sarah Rauscher
The point mutation N642H of the signal transducer and activator of transcription 5B (STAT5B) protein is associated with aggressive and drug-resistant forms of leukemia. This mutation is thought to promote cancer due to hyperactivation of STAT5B caused by increased stability of the active, parallel dimer state. However, the molecular mechanism leading to this stabilization is not well understood as there is currently no structure of the parallel dimer. To investigate the mutation's mechanism of action, we conducted extensive all-atom molecular dynamics simulations of multiple oligomeric forms of both STAT5B and STAT5B(N642H), including a model for the parallel dimer. The N642H mutation directly affects the hydrogen bonding network within the phosphotyrosine (pY)-binding pocket of the parallel dimer, enhancing the pY-binding interaction. The simulations indicate that apo STAT5B is highly flexible, exploring a diverse conformational space. In contrast, apo STAT5B(N642H) accesses two distinct conformational states, one of which resembles the conformation of the parallel dimer. The simulation predictions of the effects of the mutation on structure and dynamics are supported by the results of hydrogen-deuterium exchange (HDX) mass spectrometry measurements carried out on STAT5B and STAT5B(N642H) in which a phosphopeptide was used to mimic the effects of parallel dimerization on the SH2 domain. The molecular-level information uncovered in this work contributes to our understanding of STAT5B hyperactivation by the N642H mutation and could help pave the way for novel therapeutic strategies targeting this mutation.
{"title":"The structural influence of the oncogenic driver mutation N642H in the STAT5B SH2 domain","authors":"Liam Haas-Neill, Deniz Meneksedag-Erol, Ayesha Chaudhry, Masha Novoselova, Qirat F. Ashraf, Elvin D. de Araujo, Derek J. Wilson, Sarah Rauscher","doi":"10.1101/2024.09.09.612134","DOIUrl":"https://doi.org/10.1101/2024.09.09.612134","url":null,"abstract":"The point mutation N642H of the signal transducer and activator of transcription 5B (STAT5B) protein is associated with aggressive and drug-resistant forms of leukemia. This mutation is thought to promote cancer due to hyperactivation of STAT5B caused by increased stability of the active, parallel dimer state. However, the molecular mechanism leading to this stabilization is not well understood as there is currently no structure of the parallel dimer. To investigate the mutation's mechanism of action, we conducted extensive all-atom molecular dynamics simulations of multiple oligomeric forms of both STAT5B and STAT5B(N642H), including a model for the parallel dimer. The N642H mutation directly affects the hydrogen bonding network within the phosphotyrosine (pY)-binding pocket of the parallel dimer, enhancing the pY-binding interaction. The simulations indicate that apo STAT5B is highly flexible, exploring a diverse conformational space. In contrast, apo STAT5B(N642H) accesses two distinct conformational states, one of which resembles the conformation of the parallel dimer. The simulation predictions of the effects of the mutation on structure and dynamics are supported by the results of hydrogen-deuterium exchange (HDX) mass spectrometry measurements carried out on STAT5B and STAT5B(N642H) in which a phosphopeptide was used to mimic the effects of parallel dimerization on the SH2 domain. The molecular-level information uncovered in this work contributes to our understanding of STAT5B hyperactivation by the N642H mutation and could help pave the way for novel therapeutic strategies targeting this mutation.","PeriodicalId":501048,"journal":{"name":"bioRxiv - Biophysics","volume":"129 1","pages":""},"PeriodicalIF":0.0,"publicationDate":"2024-09-10","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142178219","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 : 2024-09-09DOI: 10.1101/2024.09.09.612070
Pooja, Pradipta Bandyopadhyay
Computational protein science has made substantial headway, but accurately predicting the functional effects of mutation in Calcium-binding proteins (CBPs) on Ca2+ binding affinity proves obscure. The complexity lies in the fact that only sequence features or structural information individually offer an incomplete picture on their own. To triumph over this adversity, we introduce a pioneering framework that effortlessly integrates protein sequence evolution information, structural characteristics, and Ca2+ binding interaction properties into a machine learning algorithm. This synthesis has been carried out poised to significantly enhance accuracy and precision in the prediction of the Ca2+ binding affinity towards CBP variants. In our study, we have developed a Ca2+ binding affinity prediction model for various mutants of cardiac Troponin-C protein, to uncover the molecular determinants that contribute binding affinity across protein variants. Our method combines state-of-the-art practices, including a physics-based approach that uses relative binding free-energy (BFE) calculations to assess mutations with implicit polarization. Additionally, it incorporates the impact of evolutionary factors on protein mutations through a theoretical deep mutational scan using a statistical probability model. Support Vector Regression (SVR) algorithms have been used to predict Ca2+ binding affinity based on sequence information, structural properties, and interactions of water molecules with Ca2+ in the EF-hand loop. Our model demonstrates high accuracy and can potentially be generalized for other calcium-binding proteins to predict the effects of point mutations on Ca2+ binding affinity for CBPs.
{"title":"Calcium Binding Affinity in the Mutational Landscape of Troponin-C: Free Energy Calculation, Co-evolution modeling and Machine Learning","authors":"Pooja, Pradipta Bandyopadhyay","doi":"10.1101/2024.09.09.612070","DOIUrl":"https://doi.org/10.1101/2024.09.09.612070","url":null,"abstract":"Computational protein science has made substantial headway, but accurately predicting the functional effects of mutation in Calcium-binding proteins (CBPs) on Ca2+ binding affinity proves obscure. The complexity lies in the fact that only sequence features or structural information individually offer an incomplete picture on their own. To triumph over this adversity, we introduce a pioneering framework that effortlessly integrates protein sequence evolution information, structural characteristics, and Ca2+ binding interaction properties into a machine learning algorithm. This synthesis has been carried out poised to significantly enhance accuracy and precision in the prediction of the Ca2+ binding affinity towards CBP variants. In our study, we have developed a Ca2+ binding affinity prediction model for various mutants of cardiac Troponin-C protein, to uncover the molecular determinants that contribute binding affinity across protein variants. Our method combines state-of-the-art practices, including a physics-based approach that uses relative binding free-energy (BFE) calculations to assess mutations with implicit polarization. Additionally, it incorporates the impact of evolutionary factors on protein mutations through a theoretical deep mutational scan using a statistical probability model. Support Vector Regression (SVR) algorithms have been used to predict Ca2+ binding affinity based on sequence information, structural properties, and interactions of water molecules with Ca2+ in the EF-hand loop. Our model demonstrates high accuracy and can potentially be generalized for other calcium-binding proteins to predict the effects of point mutations on Ca2+ binding affinity for CBPs.","PeriodicalId":501048,"journal":{"name":"bioRxiv - Biophysics","volume":"10 1","pages":""},"PeriodicalIF":0.0,"publicationDate":"2024-09-09","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142178213","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 : 2024-09-09DOI: 10.1101/2024.09.09.612093
Aldrex Munsayac, Wellington C Leite, Jesse B Hopkins, Ian Hall, Hugh M O'Neill, Sarah C Keane
The structures of RNA:RNA complexes regulate many biological processes. Despite their importance, protein-free RNA:RNA complexes represent a tiny fraction of experimentally-determined structures. Here, we describe a joint small-angle X-ray and neutron scattering (SAXS/SANS) approach to structurally interrogate conformational changes in a model RNA:RNA complex. Using SAXS, we measured the solution structures of the individual RNAs in their free state and of the overall RNA:RNA complex. With SANS, we demonstrate, as a proof-of-principle, that isotope labeling and contrast matching (CM) can be combined to probe the bound state structure of an RNA within a selectively deuterated RNA:RNA complex. Furthermore, we show that experimental scattering data can validate and improve predicted AlphaFold 3 RNA:RNA complex structures to reflect its solution structure. Our work demonstrates that in silico modeling, SAXS, and CM-SANS can be used in concert to directly analyze conformational changes within RNAs when in complex, enhancing our understanding of RNA structure in functional assemblies.
{"title":"Selective deuteration of an RNA:RNA complex for structural analysis using small-angle scattering","authors":"Aldrex Munsayac, Wellington C Leite, Jesse B Hopkins, Ian Hall, Hugh M O'Neill, Sarah C Keane","doi":"10.1101/2024.09.09.612093","DOIUrl":"https://doi.org/10.1101/2024.09.09.612093","url":null,"abstract":"The structures of RNA:RNA complexes regulate many biological processes. Despite their importance, protein-free RNA:RNA complexes represent a tiny fraction of experimentally-determined structures. Here, we describe a joint small-angle X-ray and neutron scattering (SAXS/SANS) approach to structurally interrogate conformational changes in a model RNA:RNA complex. Using SAXS, we measured the solution structures of the individual RNAs in their free state and of the overall RNA:RNA complex. With SANS, we demonstrate, as a proof-of-principle, that isotope labeling and contrast matching (CM) can be combined to probe the bound state structure of an RNA within a selectively deuterated RNA:RNA complex. Furthermore, we show that experimental scattering data can validate and improve predicted AlphaFold 3 RNA:RNA complex structures to reflect its solution structure. Our work demonstrates that in silico modeling, SAXS, and CM-SANS can be used in concert to directly analyze conformational changes within RNAs when in complex, enhancing our understanding of RNA structure in functional assemblies.","PeriodicalId":501048,"journal":{"name":"bioRxiv - Biophysics","volume":"152 1","pages":""},"PeriodicalIF":0.0,"publicationDate":"2024-09-09","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142178214","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 : 2024-09-08DOI: 10.1101/2024.09.07.611807
Haley B. Obenshain, Isaias Zarate, Olivia Hedman-Manzano, Jared Goderich, Sungho Lee, Bryant A. Lopez, Emma Varela, Ga-Young Kelly Suh, Douglas A. Pace, Siavash Ahrar
Investigating aquatic microorganisms' swimming and feeding behaviors under well-controlled conditions is of great interest across multiple disciplines. Thus, broader access to resources that enable these investigations is desirable. Given the organisms' microscopic dimensions, an ideal system should combine microscopy to visualize and fluidics to control and modulate their environments. We report an integrated device (Aquavert) that combines DIY microscopy and microfluidics for biomechanical investigations of marine microorganisms, emphasizing vertical swimming. The DIY microscope was developed for modularity, and imaging chambers were secured in vertical orientations (either in portrait or landscape mode). Fluid channels were used to introduce flow and fluid segmentation while remaining upright. Fluid segmentation established two distinct environments (e.g., with and without algae) in neighboring regions inside a chamber. System application with multiple marine larvae (sand dollars, sea urchins, and starfish) and introduction of unicellular algae were demonstrated. Finally, the device's capabilities were extended to fluorescence imaging to visualize tracer beads. The role of gravity is often ignored in conventional plate or microfluidic experiments. Beyond the current application, Aquavert enables investigations of the behavior and physiology of microorganisms where the role of gravity is critical.
{"title":"Aquavert: Imaging and Microfluidics for Vertical Swimming of Microorganisms","authors":"Haley B. Obenshain, Isaias Zarate, Olivia Hedman-Manzano, Jared Goderich, Sungho Lee, Bryant A. Lopez, Emma Varela, Ga-Young Kelly Suh, Douglas A. Pace, Siavash Ahrar","doi":"10.1101/2024.09.07.611807","DOIUrl":"https://doi.org/10.1101/2024.09.07.611807","url":null,"abstract":"Investigating aquatic microorganisms' swimming and feeding behaviors under well-controlled conditions is of great interest across multiple disciplines. Thus, broader access to resources that enable these investigations is desirable. Given the organisms' microscopic dimensions, an ideal system should combine microscopy to visualize and fluidics to control and modulate their environments. We report an integrated device (Aquavert) that combines DIY microscopy and microfluidics for biomechanical investigations of marine microorganisms, emphasizing vertical swimming. The DIY microscope was developed for modularity, and imaging chambers were secured in vertical orientations (either in portrait or landscape mode). Fluid channels were used to introduce flow and fluid segmentation while remaining upright. Fluid segmentation established two distinct environments (e.g., with and without algae) in neighboring regions inside a chamber. System application with multiple marine larvae (sand dollars, sea urchins, and starfish) and introduction of unicellular algae were demonstrated. Finally, the device's capabilities were extended to fluorescence imaging to visualize tracer beads. The role of gravity is often ignored in conventional plate or microfluidic experiments. Beyond the current application, Aquavert enables investigations of the behavior and physiology of microorganisms where the role of gravity is critical.","PeriodicalId":501048,"journal":{"name":"bioRxiv - Biophysics","volume":"80 1","pages":""},"PeriodicalIF":0.0,"publicationDate":"2024-09-08","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142178248","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 : 2024-09-08DOI: 10.1101/2024.09.03.611099
Paola Bardetti, Felix Barber, Enrique R Rojas
The bacillus - or rod - is a pervasive cellular morphology among bacteria. Rod-shaped cells elongate without widening by reinforcing their cell wall anisotropically to prevent turgor pressure from inflating cell width. Here, we demonstrate that a constrictive force is also essential for avoiding pressure-driven widening in Gram-positive bacteria. Specifically, super-resolution measurements of the nonlinear mechanical properties of the cell wall revealed that across a range of turgor pressure cell elongation directly causes width constriction, similar to a "finger trap" toy. As predicted by theory, this property depends on cell-wall anisotropy and is precisely correlated with the cell's ability to maintain a rod shape. Furthermore, the acute non-linearities in the dependence between cell length and width deformation result in a negative-feedback mechanism that confers cell-width homeostasis. That is, the Gram-positive cell wall is a "smart material" whose exotic mechanical properties are exquisitely adapted to execute cellular morphogenesis.
{"title":"Non-linear stress-softening of the bacterial cell wall confers cell shape homeostasis","authors":"Paola Bardetti, Felix Barber, Enrique R Rojas","doi":"10.1101/2024.09.03.611099","DOIUrl":"https://doi.org/10.1101/2024.09.03.611099","url":null,"abstract":"The bacillus - or rod - is a pervasive cellular morphology among bacteria. Rod-shaped cells elongate without widening by reinforcing their cell wall anisotropically to prevent turgor pressure from inflating cell width. Here, we demonstrate that a constrictive force is also essential for avoiding pressure-driven widening in Gram-positive bacteria. Specifically, super-resolution measurements of the nonlinear mechanical properties of the cell wall revealed that across a range of turgor pressure cell elongation directly causes width constriction, similar to a \"finger trap\" toy. As predicted by theory, this property depends on cell-wall anisotropy and is precisely correlated with the cell's ability to maintain a rod shape. Furthermore, the acute non-linearities in the dependence between cell length and width deformation result in a negative-feedback mechanism that confers cell-width homeostasis. That is, the Gram-positive cell wall is a \"smart material\" whose exotic mechanical properties are exquisitely adapted to execute cellular morphogenesis.","PeriodicalId":501048,"journal":{"name":"bioRxiv - Biophysics","volume":"18 1","pages":""},"PeriodicalIF":0.0,"publicationDate":"2024-09-08","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142178250","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 : 2024-09-08DOI: 10.1101/2024.09.08.611783
Nikhil Desai, Eric Lauga
The paranasal sinuses are a group of hollow spaces within the human skull, surrounding the nose. They are lined with an epithelium that contains mucus-producing cells and tiny hairlike active appendages called cilia. The cilia beat constantly to sweep mucus out of the sinus into the nasal cavity, thus maintaining a clean mucus layer within the sinuses. This process, called mucociliary clearance, is essential for a healthy nasal environment and disruption in mucus clearance leads to diseases such as chronic rhinosinusitis, specifically in the maxillary sinuses, which are the largest of the paranasal sinuses. We present here a continuum mathematical model of mucociliary clearance inside the human maxillary sinus. Using a combination of analysis and computations, we study the flow of a thin fluid film inside a fluid-producing cavity lined with an active surface: fluid is continuously produced by a wall-normal flux in the cavity and then is swept out, against gravity, due to an effective tangential flow induced by the cilia. We show that a steady layer of mucus develops over the cavity surface only when the rate of ciliary clearance exceeds a threshold, which itself depends on the rate of mucus production. We then use a scaling analysis, which highlights the competition between gravitational retention and cilia-driven drainage of mucus, to rationalise our computational results. We discuss the biological relevance of our findings, noting that measurements of mucus production and clearance rates in healthy sinuses fall within our predicted regime of steady-state mucus layer development.
{"title":"Modelling mucus clearance in sinuses: thin-film flow inside a fluid-producing cavity lined with an active surface","authors":"Nikhil Desai, Eric Lauga","doi":"10.1101/2024.09.08.611783","DOIUrl":"https://doi.org/10.1101/2024.09.08.611783","url":null,"abstract":"The paranasal sinuses are a group of hollow spaces within the human skull, surrounding the nose. They are lined with an epithelium that contains mucus-producing cells and tiny hairlike active appendages called cilia. The cilia beat constantly to sweep mucus out of the sinus into the nasal cavity, thus maintaining a clean mucus layer within the sinuses. This process, called mucociliary clearance, is essential for a healthy nasal environment and disruption in mucus clearance leads to diseases such as chronic rhinosinusitis, specifically in the maxillary sinuses, which are the largest of the paranasal sinuses. We present here a continuum mathematical model of mucociliary clearance inside the human maxillary sinus. Using a combination of analysis and computations, we study the flow of a thin fluid film inside a fluid-producing cavity lined with an active surface: fluid is continuously produced by a wall-normal flux in the cavity and then is swept out, against gravity, due to an effective tangential flow induced by the cilia. We show that a steady layer of mucus develops over the cavity surface only when the rate of ciliary clearance exceeds a threshold, which itself depends on the rate of mucus production. We then use a scaling analysis, which highlights the competition between gravitational retention and cilia-driven drainage of mucus, to rationalise our computational results. We discuss the biological relevance of our findings, noting that measurements of mucus production and clearance rates in healthy sinuses fall within our predicted regime of steady-state mucus layer development.","PeriodicalId":501048,"journal":{"name":"bioRxiv - Biophysics","volume":"40 1","pages":""},"PeriodicalIF":0.0,"publicationDate":"2024-09-08","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142178245","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 : 2024-09-08DOI: 10.1101/2024.09.05.611539
Robert C. Monsen, T. Michael Sabo, Robert D. Gray, Jesse B. Hopkins, Jonathan B. Chaires
Time-resolved small-angle X-ray experiments (TR-SAXS) are reported here that capture and quantify a previously unknown rapid collapse of the unfolded oligonucleotide as an early step in G4 folding of hybrid 1 and hybrid 2 telomeric G-quadruplex structures. The rapid collapse, initiated by a pH jump, is characterized by an exponential decrease in the radius of gyration from 20.6 to 12.6 Å. The collapse is monophasic and is complete in less than 600 ms. Additional hand-mixing pH-jump kinetic studies show that slower kinetic steps follow the collapse. The folded and unfolded states at equilibrium were further characterized by SAXS studies and other biophysical tools, to show that G4 unfolding was complete at alkaline pH, but not in LiCl solution as is often claimed. The SAXS Ensemble Optimization Method (EOM) analysis reveals models of the unfolded state as a dynamic ensemble of flexible oligonucleotide chains with a variety of transient hairpin structures. These results suggest a G4 folding pathway in which a rapid collapse, analogous to molten globule formation seen in proteins, is followed by a confined conformational search within the collapsed particle to form the native contacts ultimately found in the stable folded form.
本文报告的时间分辨小角 X 射线实验（TR-SAXS）捕获并量化了之前未知的未折叠寡核苷酸的快速塌缩，这是杂交 1 号和杂交 2 号端粒 G-四重结构 G4 折叠的早期步骤。由 pH 值跃迁引发的快速塌缩以回旋半径从 20.6 Å 到 12.6 Å 的指数下降为特征。其他手工混合 pH 值跳跃动力学研究表明，塌缩后会出现较慢的动力学步骤。通过 SAXS 研究和其他生物物理工具对平衡状态下的折叠和展开状态进行了进一步的表征，结果表明 G4 在碱性 pH 值下完全展开，而不是像通常所说的那样在氯化锂溶液中完全展开。SAXS 组合优化法（EOM）分析揭示了折叠状态的模型，即具有各种瞬时发夹结构的柔性寡核苷酸链的动态组合。这些结果表明了一种 G4 折叠路径，其中的快速塌缩类似于蛋白质中的熔融球形成，随后在塌缩颗粒内进行封闭的构象搜索，以形成最终在稳定折叠形式中发现的原生接触。
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