scholarly journals A Bacterial Surface Layer Protein Exploits Multi-step Crystallization for Rapid Self-assembly

2019 ◽  
Author(s):  
Jonathan Herrmann ◽  
Po-Nan Li ◽  
Fatemeh Jabbarpour ◽  
Anson C. K. Chan ◽  
Ivan Rajkovic ◽  
...  
Keyword(s):  
Surface Layer ◽  
Self Assembly ◽  
Molecular Mechanisms ◽  
Protein Crystallization ◽  
Crystal Nucleation ◽  
Time Resolved ◽  
Bacterial Surface ◽  
Surface Layer Protein

AbstractSurface layers (S-layers) are crystalline protein coats surrounding microbial cells. S-layer proteins (SLPs) regulate their extracellular self-assembly by crystallizing when exposed to an environmental trigger. However, molecular mechanisms governing rapid protein crystallization in vivo or in vitro are largely unknown. Here, we demonstrate that the C. crescentus SLP readily crystallizes into sheets in vitro via a calcium-triggered multi-step assembly pathway. This pathway involves two domains serving distinct functions in assembly. The C-terminal crystallization domain forms the physiological 2D crystal lattice, but full-length protein crystallizes multiple orders of magnitude faster due to the N-terminal nucleation domain. Observing crystallization using time-resolved electron cryo-microscopy (Cryo-EM) reveals a crystalline intermediate wherein N-terminal nucleation domains exhibit motional dynamics with respect to rigid lattice-forming crystallization domains. Dynamic flexibility between the two domains rationalizes efficient S-layer crystal nucleation on the curved cellular surface. Rate enhancement of protein crystallization by a discrete nucleation domain may enable engineering of kinetically controllable self-assembling 2D macromolecular nanomaterials.Significance StatementMany microbes assemble a crystalline protein layer on their outer surface as an additional barrier and communication platform between the cell and its environment. Surface layer proteins efficiently crystallize to continuously coat the cell and this trait has been utilized to design functional macromolecular nanomaterials. Here, we report that rapid crystallization of a bacterial surface layer protein occurs through a multi-step pathway involving a crystalline intermediate. Upon calcium-binding, sequential changes occur in the structure and arrangement of the protein, which are captured by time-resolved small angle x-ray scattering and transmission electron cryo-microscopy. We demonstrate that a specific domain is responsible for enhancing the rate of self-assembly, unveiling possible evolutionary mechanisms to enhance the kinetics of 2D protein crystallization in vivo.

scholarly journals A bacterial surface layer protein exploits multistep crystallization for rapid self-assembly

2019 ◽  
Vol 117 (1) ◽  
pp. 388-394 ◽  
Author(s):  
Jonathan Herrmann ◽  
Po-Nan Li ◽  
Fatemeh Jabbarpour ◽  
Anson C. K. Chan ◽  
Ivan Rajkovic ◽  
...  
Keyword(s):  
Self Assembly ◽  
Time Course ◽  
Molecular Mechanisms ◽  
Caulobacter Crescentus ◽  
Protein Crystallization ◽  
Crystal Nucleation ◽  
Bacterial Surface ◽  
Multiple Orders

Surface layers (S-layers) are crystalline protein coats surrounding microbial cells. S-layer proteins (SLPs) regulate their extracellular self-assembly by crystallizing when exposed to an environmental trigger. However, molecular mechanisms governing rapid protein crystallization in vivo or in vitro are largely unknown. Here, we demonstrate that theCaulobacter crescentusSLP readily crystallizes into sheets in vitro via a calcium-triggered multistep assembly pathway. This pathway involves 2 domains serving distinct functions in assembly. The C-terminal crystallization domain forms the physiological 2-dimensional (2D) crystal lattice, but full-length protein crystallizes multiple orders of magnitude faster due to the N-terminal nucleation domain. Observing crystallization using a time course of electron cryo-microscopy (Cryo-EM) imaging reveals a crystalline intermediate wherein N-terminal nucleation domains exhibit motional dynamics with respect to rigid lattice-forming crystallization domains. Dynamic flexibility between the 2 domains rationalizes efficient S-layer crystal nucleation on the curved cellular surface. Rate enhancement of protein crystallization by a discrete nucleation domain may enable engineering of kinetically controllable self-assembling 2D macromolecular nanomaterials.

scholarly journals Continuous, Topologically Guided Protein Crystallization Controls Bacterial Surface Layer Self-Assembly

2019 ◽  
Author(s):  
Colin J. Comerci ◽  
Jonathan Herrmann ◽  
Joshua Yoon ◽  
Fatemeh Jabbarpour ◽  
Xiaofeng Zhou ◽  
...  
Keyword(s):  
Surface Layer ◽  
Cell Surface ◽  
Self Assembly ◽  
Cell Envelope ◽  
Protein Crystallization ◽  
Bacterial Surface ◽  
Cell Surface Structures ◽  
Complicated Topology ◽  
Layer Assembly

AbstractBacteria assemble the cell envelope using localized enzymes to account for growth and division of a topologically complicated surface1–3. However, a regulatory pathway has not been identified for assembly and maintenance of the surface layer (S-layer), a 2D crystalline protein coat surrounding the curved 3D surface of a variety of bacteria4,5. By specifically labeling, imaging, and tracking native and purified RsaA, the S-layer protein (SLP) fromC. crescentus, we show that protein self-assembly alone is sufficient to assemble and maintain the S-layerin vivo. By monitoring the location of newly produced S-layer on the surface of living bacteria, we find that S-layer assembly occurs independently of the site of RsaA secretion and that localized production of new cell wall surface area alone is insufficient to explain S-layer assembly patterns. When the cell surface is devoid of a pre-existing S-layer, the location of S-layer assembly depends on the nucleation characteristics of SLP crystals, which grow by capturing RsaA molecules freely diffusing on the outer bacterial surface. Based on these observations, we propose a model of S-layer assembly whereby RsaA monomers are secreted randomly and diffuse on the lipopolysaccharide (LPS) outer membrane until incorporated into growing 2D S-layer crystals. The complicated topology of the cell surface enables formation of defects, gaps, and grain boundaries within the S-layer lattice, thereby guiding the location of S-layer assembly without enzymatic assistance. This unsupervised mechanism poses unique challenges and advantages for designing treatments targeting cell surface structures or utilizing S-layers as self-assembling macromolecular nanomaterials. As an evolutionary driver, 2D protein self-assembly rationalizes the exceptional S-layer subunit sequence and species diversity6.

scholarly journals Cotranslational folding allows misfolding-prone proteins to circumvent deep kinetic traps

2020 ◽  
Vol 117 (3) ◽  
pp. 1485-1495 ◽  
Author(s):  
Amir Bitran ◽  
William M. Jacobs ◽  
Xiadi Zhai ◽  
Eugene Shakhnovich
Keyword(s):  
Self Assembly ◽  
Molecular Mechanisms ◽  
Folding Kinetics ◽  
Rare Codons ◽  
Cotranslational Folding ◽  
Nascent Chain ◽  
Kinetic Traps ◽  
Optimal Window

Many large proteins suffer from slow or inefficient folding in vitro. It has long been known that this problem can be alleviated in vivo if proteins start folding cotranslationally. However, the molecular mechanisms underlying this improvement have not been well established. To address this question, we use an all-atom simulation-based algorithm to compute the folding properties of various large protein domains as a function of nascent chain length. We find that for certain proteins, there exists a narrow window of lengths that confers both thermodynamic stability and fast folding kinetics. Beyond these lengths, folding is drastically slowed by nonnative interactions involving C-terminal residues. Thus, cotranslational folding is predicted to be beneficial because it allows proteins to take advantage of this optimal window of lengths and thus avoid kinetic traps. Interestingly, many of these proteins’ sequences contain conserved rare codons that may slow down synthesis at this optimal window, suggesting that synthesis rates may be evolutionarily tuned to optimize folding. Using kinetic modeling, we show that under certain conditions, such a slowdown indeed improves cotranslational folding efficiency by giving these nascent chains more time to fold. In contrast, other proteins are predicted not to benefit from cotranslational folding due to a lack of significant nonnative interactions, and indeed these proteins’ sequences lack conserved C-terminal rare codons. Together, these results shed light on the factors that promote proper protein folding in the cell and how biomolecular self-assembly may be optimized evolutionarily.

scholarly journals Spontaneous self-assembly of pathogenic huntingtin exon 1 protein into amyloid structures

2014 ◽  
Vol 56 ◽  
pp. 167-180 ◽  
Author(s):  
Philipp Trepte ◽  
Nadine Strempel ◽  
Erich E. Wanker
Keyword(s):  
Self Assembly ◽  
Molecular Mechanisms ◽  
Critical Role ◽  
Cytoplasmic Inclusion ◽  
Spinocerebellar Ataxia Type ◽  
Neuronal Processes ◽  
Exon 1 ◽  
Novel Therapeutic Strategies

PolyQ (polyglutamine) diseases such as HD (Huntington's disease) or SCA1 (spinocerebellar ataxia type 1) are neurodegenerative disorders caused by abnormally elongated polyQ tracts in human proteins. PolyQ expansions promote misfolding and aggregation of disease-causing proteins, leading to the appearance of nuclear and cytoplasmic inclusion bodies in patient neurons. Several lines of experimental evidence indicate that this process is critical for disease pathogenesis. However, the molecular mechanisms underlying spontaneous polyQ-containing aggregate formation and the perturbation of neuronal processes are still largely unclear. The present chapter reviews the current literature regarding misfolding and aggregation of polyQ-containing disease proteins. We specifically focus on studies that have investigated the amyloidogenesis of polyQ-containing HTTex1 (huntingtin exon 1) fragments. These protein fragments are disease-relevant and play a critical role in HD pathogenesis. We outline potential mechanisms behind mutant HTTex1 aggregation and toxicity, as well as proteins and small molecules that can modify HTTex1 amyloidogenesis in vitro and in vivo. The potential implications of such studies for the development of novel therapeutic strategies are discussed.

scholarly journals Bacillus anthracis SlaQ Promotes S-Layer Protein Assembly

2015 ◽  
Vol 197 (19) ◽  
pp. 3216-3227 ◽  
Author(s):  
Sao-Mai Nguyen-Mau ◽  
So-Young Oh ◽  
Daphne I. Schneewind ◽  
Dominique Missiakas ◽  
Olaf Schneewind
Keyword(s):  
Bacillus Anthracis ◽  
Self Assembly ◽  
Protein Assembly ◽  
Crystalline Lattice ◽  
Cell Wall Polysaccharide ◽  
Cytoplasmic Protein ◽  
Content Type ◽  
Bacterial Surface

ABSTRACTBacillus anthracisvegetative forms assemble an S-layer comprised of two S-layer proteins, Sap and EA1. A hallmark of S-layer proteins are their C-terminal crystallization domains, which assemble into a crystalline lattice once these polypeptides are deposited on the bacterial surface via association between their N-terminal S-layer homology domains and the secondary cell wall polysaccharide. Here we show thatslaQ, encoding a small cytoplasmic protein conserved among pathogenic bacilli elaborating S-layers, is required for the efficient secretion and assembly of Sap and EA1. S-layer protein precursors cosediment with SlaQ, and SlaQ appears to facilitate Sap assembly. Purified SlaQ polymerizes and when mixed with purified Sap promotes thein vitroformation of tubular S-layer structures. A model is discussed whereby SlaQ, in conjunction with S-layer secretion factors SecA2 and SlaP, promotes localized secretion and S-layer assembly inB. anthracis.IMPORTANCES-layer proteins are endowed with the propensity for self-assembly into crystalline arrays. Factors promoting S-layer protein assembly have heretofore not been reported. We identifiedBacillus anthracisSlaQ, a small cytoplasmic protein that facilitates S-layer protein assemblyin vivoandin vitro.

scholarly journals Topologically-guided continuous protein crystallization controls bacterial surface layer self-assembly

2019 ◽  
Vol 10 (1) ◽  
Author(s):  
Colin J. Comerci ◽  
Jonathan Herrmann ◽  
Joshua Yoon ◽  
Fatemeh Jabbarpour ◽  
Xiaofeng Zhou ◽  
...  
Keyword(s):  
Surface Layer ◽  
Self Assembly ◽  
Protein Crystallization ◽  
Bacterial Surface

Monte Carlo study of the molecular mechanisms of surface-layer protein self-assembly

2011 ◽  
Vol 134 (12) ◽  
pp. 125103 ◽  
Author(s):  
Christine Horejs ◽  
Mithun K. Mitra ◽  
Dietmar Pum ◽  
Uwe B. Sleytr ◽  
Murugappan Muthukumar
Keyword(s):  
Monte Carlo ◽  
Surface Layer ◽  
Self Assembly ◽  
Molecular Mechanisms ◽  
Monte Carlo Study ◽  
Surface Layer Protein

scholarly journals Continuous, Topologically Guided Protein Crystallization Drives Self-Assembly of a Bacterial Surface Layer

2020 ◽  
Vol 118 (3) ◽  
pp. 201a-202a
Author(s):  
Colin J. Comerci ◽  
Jonathan Herrmann ◽  
Joshua Yoon ◽  
Fatemeh Jabbarpour ◽  
Xiaofeng Zhou ◽  
...  
Keyword(s):  
Surface Layer ◽  
Self Assembly ◽  
Protein Crystallization ◽  
Bacterial Surface

In vitro and in vivo polysaccharides assembly: ultrastructural and cytochemical study of growing plant cell wall components.

1978 ◽  
Vol 36 (2) ◽  
pp. 434-435
Author(s):  
D. Reis ◽  
B. Vian ◽  
J. C. Roland
Keyword(s):  
Cell Wall ◽  
Self Assembly ◽  
Plant Cell Wall ◽  
Higher Plants ◽  
Three Dimensional ◽  
Cytochemical Study ◽  
Wall Components

Wall morphogenesis in higher plants is a problem still open to controversy. Until now the possibility of a transmembrane control and the involvement of microtubules were mostly envisaged. Self-assembly processes have been observed in the case of walls of Chlamydomonas and bacteria. Spontaneous gelling interactions between xanthan and galactomannan from Ceratonia have been analyzed very recently. The present work provides indications that some processes of spontaneous aggregation could occur in higher plants during the formation and expansion of cell wall.Observations were performed on hypocotyl of mung bean (Phaseolus aureus) for which growth characteristics and wall composition have been previously defined.In situ, the walls of actively growing cells (primary walls) show an ordered three-dimensional organization (fig. 1). The wall is typically polylamellate with multifibrillar layers alternately transverse and longitudinal. Between these layers intermediate strata exist in which the orientation of microfibrils progressively rotates. Thus a progressive change in the morphogenetic activity occurs.

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