Intrinsic aggregation propensity of the CsgB nucleator protein is crucial for curli fiber formation

2016 ◽  
Vol 195 (2) ◽  
pp. 179-189 ◽  
Author(s):  
Nikolaos N. Louros ◽  
Georgios M.P. Bolas ◽  
Paraskevi L. Tsiolaki ◽  
Stavros J. Hamodrakas ◽  
Vassiliki A. Iconomidou
Keyword(s):  
Fiber Formation ◽  
Aggregation Propensity ◽  
Curli Fiber

scholarly journals Structure-Function Analysis of the Curli Accessory Protein CsgE Defines Surfaces Essential for Coordinating Amyloid Fiber Formation

mBio ◽  
2018 ◽  
Vol 9 (4) ◽  
Author(s):  
Roger D. Klein ◽  
Qin Shu ◽  
Zachary T. Cusumano ◽  
Kanna Nagamatsu ◽  
Nathaniel C. Gualberto ◽  
...  
Keyword(s):  
Outer Membrane ◽  
Fiber Formation ◽  
Single Amino Acid ◽  
Amyloid Fibers ◽  
Membrane Pore ◽  
Curli Fiber ◽  
Functional Amyloids ◽  
Positively Charged ◽  
Biofilm Matrix

ABSTRACT Curli amyloid fibers are produced as part of the extracellular biofilm matrix and are composed primarily of the major structural subunit CsgA. The CsgE chaperone facilitates the secretion of CsgA through CsgG by forming a cap at the base of the nonameric CsgG outer membrane pore. We elucidated a series of finely tuned nonpolar and charge-charge interactions that facilitate the oligomerization of CsgE and its ability to transport unfolded CsgA to CsgG for translocation. CsgE oligomerization in vitro is temperature dependent and is disrupted by mutations in the W48 and F79 residues. Using nuclear magnetic resonance (NMR), we identified two regions of CsgE involved in the CsgE-CsgA interaction: a head comprising a positively charged patch centered around R47 and a stem comprising a negatively charged patch containing E31 and E85. Negatively charged residues in the intrinsically disordered N- and C-terminal “tails” were not implicated in this interaction. Head and stem residues were mutated and interrogated using in vivo measurements of curli production and in vitro amyloid polymerization assays. The R47 head residue of CsgE is required for stabilization of CsgA- and CsgE-mediated curli fiber formation. Mutation of the E31 and E85 stem residues to positively charged side chains decreased CsgE-mediated curli fiber formation but increased CsgE-mediated stabilization of CsgA. No single-amino-acid substitutions in the head, stem, or tail regions affected the ability of CsgE to cap the CsgG pore as determined by a bile salt sensitivity assay. These mechanistic insights into the directed assembly of functional amyloids in extracellular biofilms elucidate possible targets for biofilm-associated bacterial infections. IMPORTANCE Curli represent a class of functional amyloid fibers produced by Escherichia coli and other Gram-negative bacteria that serve as protein scaffolds in the extracellular biofilm matrix. Despite the lack of sequence conservation among different amyloidogenic proteins, the structural and biophysical properties of functional amyloids such as curli closely resemble those of amyloids associated with several common neurodegenerative diseases. These parallels are underscored by the observation that certain proteins and chemicals can prevent amyloid formation by the major curli subunit CsgA and by alpha-synuclein, the amyloid-forming protein found in Lewy bodies during Parkinson’s disease. CsgA subunits are targeted to the CsgG outer membrane pore by CsgE prior to secretion and assembly into fibers. Here, we use biophysical, biochemical, and genetic approaches to elucidate a mechanistic understanding of CsgE function in curli biogenesis.

scholarly journals Spatial Clustering of the Curlin Secretion Lipoprotein Requires Curli Fiber Assembly

2008 ◽  
Vol 191 (2) ◽  
pp. 608-615 ◽  
Author(s):  
Elisabeth Ashman Epstein ◽  
Margeaux A. Reizian ◽  
Matthew R. Chapman
Keyword(s):  
Cell Surface ◽  
Outer Membrane ◽  
Spatial Clustering ◽  
Microscopic Analysis ◽  
Fiber Formation ◽  
Gram Negative Bacteria ◽  
Electron Microscopic ◽  
Fiber Assembly ◽  
Curli Fiber ◽  
Mutant Strains

ABSTRACT Gram-negative bacteria assemble functional amyloid surface fibers called curli. CsgB nucleates the major curli subunit protein, CsgA, into a self-propagating amyloid fiber on the cell surface. The CsgG lipoprotein is sufficient for curlin transport across the outer membrane and is hypothesized to be the central molecule of the curli fiber secretion and assembly complex. We tested the hypothesis that the curli secretion protein, CsgG, was restricted to certain areas of the cell to promote the interaction of CsgA and CsgB during curli assembly. Here, electron microscopic analysis of curli-producing strains showed that relatively few cells in the population contacted curli fibers and that curli emanated from spatially discrete points on the cell surface. Microscopic analysis revealed that CsgG was surface exposed and spatially clustered around curli fibers. CsgG localization to the outer membrane and exposure of the surface domain were not dependent on any other csg-encoded protein, but the clustering of CsgG required the csg-encoded proteins CsgE, CsgF, CsgA, and CsgB. CsgG formed stable oligomers in all the csg mutant strains, but these oligomers were distinct from the CsgG complexes assembled in wild-type cells. Finally, we found that efficient fiber assembly was required for the spatial clustering of CsgG. These results suggest a new model where curli fiber formation is spatially coordinated with the CsgG assembly apparatus.

Current Challenges and Limitations in the Studies of Intrinsically Disordered Proteins in Neurodegenerative Diseases by Computer Simulations

2020 ◽  
Vol 17 ◽  
Author(s):  
Ibrahim Yagiz Akbayrak ◽  
Sule Irem Caglayan ◽  
Zilan Ozcan ◽  
Vladimir N. Uversky ◽  
Orkid Coskuner-Weber
Keyword(s):  
Neurodegenerative Diseases ◽  
Computer Simulations ◽  
Conformational Changes ◽  
Intrinsically Disordered Proteins ◽  
Disordered Proteins ◽  
Computational Studies ◽  
Accurate Data ◽  
Aggregation Propensity ◽  
Intrinsically Disordered ◽  
Future Developments

: Experiments face challenges in the analysis of intrinsically disordered proteins in solution due to fast conformational changes and enhanced aggregation propensity. Computational studies complement experiments, being widely used in the analyses of intrinsically disordered proteins, especially those positioned at the centers of neurodegenerative diseases. However, recent investigations – including our own – revealed that computer simulations face significant challenges and limitations themselves. In this review, we introduced and discussed some of the scientific challenges and limitations of computational studies conducted on intrinsically disordered proteins. We also outlined the importance of future developments in the areas of computational chemistry and computational physics that would be needed for generating more accurate data for intrinsically disordered proteins from computer simulations. Additional theoretical strategies that can be developed are discussed herein.

Effect of Ionic Strength on the Aggregation Propensity of Aβ1-42 Peptide: An In-silico Study

2020 ◽  
Vol 14 (3) ◽  
pp. 216-226
Author(s):  
Priyanka Borah ◽  
Venkata S.K. Mattaparthi
Keyword(s):  
Diffusion Coefficient ◽  
Ionic Strength ◽  
Marked Effect ◽  
Computational Study ◽  
Conformational Dynamics ◽  
Rapid Change ◽  
Md Simulations ◽  
Radius Of Gyration ◽  
Aggregation Propensity ◽  
In Silico Study

Background: Aggregation of misfolded proteins under stress conditions in the cell might lead to several neurodegenerative disorders. Amyloid-beta (Aβ1-42) peptide, the causative agent of Alzheimer’s disease, has the propensity to fold into β-sheets under stress, forming aggregated amyloid plaques. This is influenced by factors such as pH, temperature, metal ions, mutation of residues, and ionic strength of the solution. There are several studies that have highlighted the importance of ionic strength in affecting the folding and aggregation propensity of Aβ1-42 peptide. Objective: To understand the effect of ionic strength of the solution on the aggregation propensity of Aβ1-42 peptide, using computational approaches. Materials and Methods: In this study, Molecular Dynamics (MD) simulations were performed on Aβ1-42 peptide monomer placed in (i) 0 M, (ii) 0.15 M, and (iii) 0.30 M concentration of NaCl solution. To prepare the input files for the MD simulations, we have used the Amberff99SB force field. The conformational dynamics of Aβ1-42 peptide monomer in different ionic strengths of the solutions were illustrated from the analysis of the corresponding MD trajectory using the CPPtraj tool. Results: From the MD trajectory analysis, we observe that with an increase in the ionic strength of the solution, Aβ1-42 peptide monomer shows a lesser tendency to undergo aggregation. From RMSD and SASA analysis, we noticed that Aβ1-42 peptide monomer undergoes a rapid change in conformation with an increase in the ionic strength of the solution. In addition, from the radius of gyration (Rg) analysis, we observed Aβ1-42 peptide monomer to be more compact at moderate ionic strength of the solution. Aβ1-42 peptide was also found to hold its helical secondary structure at moderate and higher ionic strengths of the solution. The diffusion coefficient of Aβ1-42 peptide monomer was also found to vary with the ionic strength of the solution. We observed a relatively higher diffusion coefficient value for Aβ1-42 peptide at moderate ionic strength of the solution. Conclusion: Our findings from this computational study highlight the marked effect of ionic strength of the solution on the conformational dynamics and aggregation propensity of Aβ1-42 peptide monomer.

A polyamide from bisacid A2 and p-phenylenediamine: Synthesis, properties, and fiber formation

1977 ◽  
Vol 21 (6) ◽  
pp. 1543-1559 ◽  
Author(s):  
Rudolph S. Lenk ◽  
James L. White ◽  
John F. Fellers
Keyword(s):  
Fiber Formation ◽  
Synthesis Properties

scholarly journals Fiber Formation and Structural Development of HBA/HNA Thermotropic Liquid Crystalline Polymer in High-Speed Melt Spinning

Polymers ◽  
2021 ◽  
Vol 13 (7) ◽  
pp. 1134
Author(s):  
Bo Seok Song ◽  
Jun Young Lee ◽  
Sun Hwa Jang ◽  
Wan-Gyu Hahm
Keyword(s):  
Mechanical Properties ◽  
Shear Rate ◽  
High Speed ◽  
Melt Spinning ◽  
Liquid Crystalline Polymer ◽  
Liquid Crystalline ◽  
Crystalline Polymer ◽  
Fiber Formation ◽  
Structural Development ◽  
Thermotropic Liquid Crystalline Polymer

High-speed melt spinning of thermotropic liquid crystalline polymer (TLCP) resin composed of 4-hydroxybenzoic acid (HBA) and 2-hydroxy-6-napthoic acid (HNA) monomers in a molar ratio of 73/27 was conducted to investigate the characteristic structure development of the fibers under industrial spinning conditions, and the obtained as-spun TLCP fibers were analyzed in detail. The tensile strength and modulus of the fibers increased with shear rate in nozzle hole, draft in spin-line and spinning temperature and exhibited the high values of approximately 1.1 and 63 GPa, respectively, comparable to those of industrial as-spun TLCP fibers, at a shear rate of 70,000 s−1 and a draft of 25. X-ray diffraction demonstrated that the mechanical properties of the fibers increased with the crystalline orientation factor (fc) and the fractions of highly oriented crystalline and non-crystalline anisotropic phases. The results of structure analysis indicated that a characteristic skin–core structure developed at high drafts (i.e., spinning velocity) and low spinning temperatures, which contributed to weakening the mechanical properties of the TLCP fibers. It is supposed that this heterogeneous structure in the cross-section of the fibers was induced by differences in the cooling rates of the skin and core of the fiber in the spin-line.

scholarly journals EpCAM promotes endosomal modulation of the cortical RhoA zone for epithelial organization

2021 ◽  
Vol 12 (1) ◽  
Author(s):  
Cécile Gaston ◽  
Simon De Beco ◽  
Bryant Doss ◽  
Meng Pan ◽  
Estelle Gauquelin ◽  
...  
Keyword(s):  
Cell Shape ◽  
Specific Protein ◽  
Cell Polarization ◽  
Stress Fiber ◽  
Fiber Formation ◽  
Endosomal Trafficking ◽  
Transient Zone ◽  
And Behavior ◽  
Fine Tune

AbstractAt the basis of cell shape and behavior, the organization of actomyosin and its ability to generate forces are widely studied. However, the precise regulation of this contractile network in space and time is unclear. Here, we study the role of the epithelial-specific protein EpCAM, a contractility modulator, in cell shape and motility. We show that EpCAM is required for stress fiber generation and front-rear polarity acquisition at the single cell level. In fact, EpCAM participates in the remodeling of a transient zone of active RhoA at the cortex of spreading epithelial cells. EpCAM and RhoA route together through the Rab35/EHD1 fast recycling pathway. This endosomal pathway spatially organizes GTP-RhoA to fine tune the activity of actomyosin resulting in polarized cell shape and development of intracellular stiffness and traction forces. Impairment of GTP-RhoA endosomal trafficking either by silencing EpCAM or by expressing Rab35/EHD1 mutants prevents proper myosin-II activity, stress fiber formation and ultimately cell polarization. Collectively, this work shows that the coupling between co-trafficking of EpCAM and RhoA, and actomyosin rearrangement is pivotal for cell spreading, and advances our understanding of how biochemical and mechanical properties promote cell plasticity.

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