Shell Subunits Research Articles

Modeling bacterial microcompartment architectures for enhanced cyanobacterial carbon fixation.

The carboxysome is a bacterial microcompartment (BMC) which plays a central role in the cyanobacterial CO2-concentrating mechanism. These proteinaceous structures consist of an outer protein shell that partitions Rubisco and carbonic anhydrase from the rest of the cytosol, thereby providing a favorable microenvironment that enhances carbon fixation. The modular nature of carboxysomal architectures makes them attractive for a variety of biotechnological applications such as carbon capture and utilization. In silico approaches, such as molecular dynamics (MD) simulations, can support future carboxysome redesign efforts by providing new spatio-temporal insights on their structure and function beyond in vivo experimental limitations. However, specific computational studies on carboxysomes are limited. Fortunately, all BMC (including the carboxysome) are highly structurally conserved which allows for practical inferences to be made between classes. Here, we review simulations on BMC architectures which shed light on (1) permeation events through the shell and (2) assembly pathways. These models predict the biophysical properties surrounding the central pore in BMC-H shell subunits, which in turn dictate the efficiency of substrate diffusion. Meanwhile, simulations on BMC assembly demonstrate that assembly pathway is largely dictated kinetically by cargo interactions while final morphology is dependent on shell factors. Overall, these findings are contextualized within the wider experimental BMC literature and framed within the opportunities for carboxysome redesign for biomanufacturing and enhanced carbon fixation.

Monatomic ions influence substrate permeation across bacterial microcompartment shells

Bacterial microcompartments (BMCs) are protein organelles consisting of an inner enzymatic core encased within a selectively permeable shell. BMC shells are modular, tractable architectures that can be repurposed with new interior enzymes for biomanufacturing purposes. The permeability of BMC shells is function-specific and regulated by biophysical properties of the shell subunits, especially its pores. We hypothesized that ions may interact with pore residues in a manner that influences the substrate permeation process. In vitro activity comparisons between native and broken BMCs demonstrated that increasing NaCl negatively affects permeation rates. Molecular dynamics simulations of the dominant shell protein (BMC-H) revealed that chloride ions preferentially occupy the positive pore, hindering substrate permeation, while sodium cations remain excluded. Overall, these results demonstrate that shell properties influence ion permeability and leverages the integration of experimental and computational techniques to improve our understanding of BMC shells towards their repurposing for biotechnological applications.

The COPII subunit MoSec24B is involved in development, pathogenicity and autophagy in the rice blast fungus.

The endoplasmic reticulum (ER) acts as the starting point of the secretory pathway, where approximately one-third of the proteins are correctly folded and modified, loaded into vesicles, and transported to the Golgi for further processing and modification. In this process, COPII vesicles are responsible for transporting cargo proteins from the ER to the Golgi. Here, we identified the inner shell subunit of COPII vesicles (MoSec24B) and explored the importance of MoSec24B in the rice blast fungus. The targeted disruption of MoSec24B led to decreased growth, reduced conidiation, restricted glycogen and lipids utilization, sensitivity to the cell wall and hypertonic stress, the failure of septin-mediated repolarization of appressorium, impaired appressorium turgor pressure, and decreased ability to infect, which resulted in reduced pathogenicity to the host plant. Furthermore, MoSec24B functions in the three mitogen-activated protein kinase (MAPK) signaling pathways by acting with MoMst50. Deletion of MoSec24B caused reduced lipidation of MoAtg8, accelerated degradation of exogenously introduced GFP-MoAtg8, and increased lipidation of MoAtg8 upon treatment with a late inhibitor of autophagy (BafA1), suggesting that MoSec24B regulates the fusion of late autophagosomes with vacuoles. Together, these results suggest that MoSec24B exerts a significant role in fungal development, the pathogenesis of filamentous fungi and autophagy.

Occurrence and stability of hetero-hexamer associations formed by β-carboxysome CcmK shell components.

The carboxysome is a bacterial micro-compartment (BMC) subtype that encapsulates enzymatic activities necessary for carbon fixation. Carboxysome shells are composed of a relatively complex cocktail of proteins, their precise number and identity being species dependent. Shell components can be classified in two structural families, the most abundant class associating as hexamers (BMC-H) that are supposed to be major players for regulating shell permeability. Up to recently, these proteins were proposed to associate as homo-oligomers. Genomic data, however, demonstrated the existence of paralogs coding for multiple shell subunits. Here, we studied cross-association compatibilities among BMC-H CcmK proteins of Synechocystis sp. PCC6803. Co-expression in Escherichia coli proved a consistent formation of hetero-hexamers combining CcmK1 and CcmK2 or, remarkably, CcmK3 and CcmK4 subunits. Unlike CcmK1/K2 hetero-hexamers, the stoichiometry of incorporation of CcmK3 in associations with CcmK4 was low. Cross-interactions implicating other combinations were weak, highlighting a structural segregation of the two groups that could relate to gene organization. Sequence analysis and structural models permitted the localization of interactions that would favor formation of CcmK3/K4 hetero-hexamers. The crystallization of these CcmK3/K4 associations conducted to the elucidation of a structure corresponding to the CcmK4 homo-hexamer. Yet, subunit exchange could not be demonstrated in vitro. Biophysical measurements showed that hetero-hexamers are thermally less stable than homo-hexamers, and impeded in forming larger assemblies. These novel findings are discussed within the context of reported data to propose a functional scenario in which minor CcmK3/K4 incorporation in shells would introduce sufficient local disorder as to allow shell remodeling necessary to adapt rapidly to environmental changes.

The Plasticity of Molecular Interactions Governs Bacterial Microcompartment Shell Assembly

Bacterial microcompartments (BMCs) are composed of an enzymatic core encapsulated by a selectively permeable protein shell that enhances catalytic efficiency. Many pathogenic bacteria derive competitive advantages from their BMC-based catabolism, implicating BMCs as drug targets. BMC shells are of interest for bioengineering due to their diverse and selective permeability properties and because they self-assemble. A complete understanding of shell composition and organization is a prerequisite for biotechnological applications. Here, we report the cryoelectron microscopy structure of a BMC shell at 3.0-Å resolution, using an image-processing strategy that allowed us to determine the previously uncharacterized structural details of the interactions formed by the BMC-TS and BMC-TD shell subunits in the context of the assembled shell. We found unexpected structural plasticity among these interactions, resulting in distinct shell populations assembled from varying numbers of the BMC-TS and BMC-TD subunits. We discuss the implications of these findings on shell assembly and function.

A New Approach for Free Vibration Analysis of Thin‐Walled Box Girder Considering Shear Lag Effect

The thin‐walled box girder (T‐WBG) is widely applied in the long‐span bridge structures during the past decades due to its lighter self‐weight and better mechanical properties. The shear lag effect (SLE), an essential aspect of T‐WBG which governs the stress and the deformation, is rather necessary to be revealed properly. The extraordinary issue of T‐WBG analysis nowadays is the SLE impact on its dynamical response to external load. This paper proposes an improved finite element method (FEM) to obtain the realistic vibration characteristics of the T‐WBG considering the SLE by theory analysis and formula derivation. Firstly, based on the classical plate and shell theory as well as beam theory, the T‐WBG was divided into shell subunit for the roof and beam subunit for web and floor, respectively. Secondly, a 3‐order polynomial which is consistent with the experiment results was adopted as the axial‐displacement interpolation function of the roof subunit, whose nodal displacements parameters were also taken as the basic. Thirdly, the nodal displacement parameters of the web subunit and floor subunit were deduced by the basic according to the principle of deflection consistency. It is shown through a numerical example that the proposed method is much more economical to achieve reasonable accuracy than traditional FEM analysis software when dealing with the free vibration problem of the T‐WBG considering the SLE. Besides, it is also observed that the natural frequency values considering the SLE have a trend of decreasing markedly in general, and the influence of SLE on higher‐order frequency is more significant than on the lower one under the boundary condition of cantilever supported, while a contrary effect under the boundary condition of simple supported.

Protein Nanotubes: Purification and Characterization of Protein Nanotubes Assembled from a Single Bacterial Microcompartment Shell Subunit (Adv. Mater. Interfaces 1/2016)

Protein Nanotubes: Purification and Characterization of Protein Nanotubes Assembled from a Single Bacterial Microcompartment Shell Subunit (Adv. Mater. Interfaces 1/2016)

Purification and Characterization of Protein Nanotubes Assembled from a Single Bacterial Microcompartment Shell Subunit

Purification and Characterization of Protein Nanotubes Assembled from a Single Bacterial Microcompartment Shell Subunit

Dynamic structural evolution of supported palladium-ceria core-shell catalysts revealed by in situ electron microscopy.

The exceptional activity for methane combustion of modular palladium–ceria core–shell subunits on silicon-functionalized alumina that was recently reported has created renewed interest in the potential of core–shell structures as catalysts. Here we report on our use of advanced ex situ and in situ electron microscopy with atomic resolution to show that the modular palladium–ceria core–shell subunits undergo structural evolution over a wide temperature range. In situ observations performed in an atmospheric gas cell within this temperature range provide real-time evidence that the palladium and ceria nanoparticle constituents of the palladium–ceria core–shell participate in a dynamical process that leads to the formation of an unanticipated structure comprised of an intimate mixture of palladium, cerium, silicon and oxygen, with very high dispersion. This finding may open new perspectives about the origin of the activity of this catalyst.

A Family of Mesocubes

It is challenging to develop a general universal procedure to fabricate mesoscale cubic structures on a large scale with different nanoscale building units. It is always desirable to tune the chemical compositions within confined arrangements without damaging the mesostructures to provide the desired physiochemical properties required by various devices/applications. Herein, we report the successful design and facile preparation of a family of mesocubes with different compositions, including (a) ZnSn(OH)6, (b) evenly distributed Zn2SnO4 and SnO2 nanoparticles, (c) hollow cubes of SnO2 nanoparticles, (d) high-ordered nanoparticles of Zn2SnO4&Sn@C; (e) SnO2@C core–shell subunits, (f) SnO2@C nanoparticle aggregates enclosed with oxidized carbon sheath, and (g) C nanobubbles, as building units, all, except ZnSn(OH)6, with the same confined arrangements of nanoparticles as building units inside the same framework of cubic mesostructures. This family of mesocubes will provide a rich pool of materials with different functional properties to meet demands in different applications and offer opportunities to evaluate fundamentals of structure–property–performance relationships. On the basis of the best of our knowledge, this family of facilely prepared mesocubes with unique combination of microsize cubes and compositions was reported for the first time, especially the carbon mesocubes formed by aggregation of carbon nanobubbles as the building subunits. Additionally, we demonstrated, for the first time, that two family members of mesocubes of Zn2SnO4&SnO2 and Zn2SnO4&Sn@C can be used as anode materials in lithium ion batteries with impressive high packing densities and superior rate performance.

Iron Acquisition in Bacillus cereus: The Roles of IlsA and Bacillibactin in Exogenous Ferritin Iron Mobilization

In host-pathogen interactions, the struggle for iron may have major consequences on the outcome of the disease. To overcome the low solubility and bio-availability of iron, bacteria have evolved multiple systems to acquire iron from various sources such as heme, hemoglobin and ferritin. The molecular basis of iron acquisition from heme and hemoglobin have been extensively studied; however, very little is known about iron acquisition from host ferritin, a 24-mer nanocage protein able to store thousands of iron atoms within its cavity. In the human opportunistic pathogen Bacillus cereus, a surface protein named IlsA (Iron-regulated leucine rich surface protein type A) binds heme, hemoglobin and ferritin in vitro and is involved in virulence. Here, we demonstrate that IlsA acts as a ferritin receptor causing ferritin aggregation on the bacterial surface. Isothermal titration calorimetry data indicate that IlsA binds several types of ferritins through direct interaction with the shell subunits. UV-vis kinetic data show a significant enhancement of iron release from ferritin in the presence of IlsA indicating for the first time that a bacterial protein might alter the stability of the ferritin iron core. Disruption of the siderophore bacillibactin production drastically reduces the ability of B. cereus to utilize ferritin for growth and results in attenuated bacterial virulence in insects. We propose a new model of iron acquisition in B. cereus that involves the binding of IlsA to host ferritin followed by siderophore assisted iron uptake. Our results highlight a possible interplay between a surface protein and a siderophore and provide new insights into host adaptation of B. cereus and general bacterial pathogenesis.

Potential energy curves via double electron-attachment calculations: Dissociation of alkali metal dimers

The recently developed method [M. Musiał, J. Chem. Phys. 136, 134111 (2012)] to study double electron attached states has been applied to the description of the ground and excited state potential energy curves of the alkali metal dimers. The method is based on the multireference coupled cluster scheme formulated within the Fock space formalism for the (2,0) sector. Due to the use of the efficient intermediate Hamiltonian formulation, the approach is free from the intruder states problem. The description of the neutral alkali metal dimers is accomplished via attaching two electrons to the corresponding doubly ionized system. This way is particularly advantageous when a closed shell molecule dissociates into open shell subunits while its doubly positive cation generates the closed shell fragments. In the current work, we generate the potential energy curves for the ground and multiple excited states of the Li2 and Na2 molecules. In all cases the potential energy curves are smooth for the entire range of interatomic distances (from the equilibrium point to the dissociation limit). Based on the calculated potential energy curves, we are able to compute spectroscopic parameters of the systems studied.

Density functional study on the electronic properties, polarizabilities, NICS values, and absorption spectra of fluorinated fullerene derivative C60F17CF3

Electronic structures, polarizabilities, NICS values, and absorption spectra of the fluorinated fullerene derivative C60F17CF3 have been systematically studied by density functional theory. The large Eg (2.39eV) between HOMO and LUMO and the strong aromatic character (with NICS −7.25ppm) of C60F17CF3 indicate it posses high stability. Further investigations show that C60F17CF3 could be excellent electron acceptors for potential photonic/photovoltaic applications in consequence of its large VIP (8.78eV). The density of states and frontier molecular orbitals are also calculated, which present that HOMOs and LUMOs are mainly distributed in the tortoise shell subunit of C60F17CF3, and the influence from F and CF3 is secondary. Our calculations shows that the absorption spectra of C60F17CF3 structure is distorted by the F and CF3 addition and the optical gaps of C60F17CF3 is red shifted relative to that of C60. The attached F and CF3 in C60F17CF3 greatly change the cage and disrupt aromatic rings, which lead to stronger aromatic character of itself than that of C60. In addition, the static linear polarizability α and first-order hyperpolarizability β0 for C60F17CF3 are significantly larger than those of C60 because of its lower symmetric structures and high delocalization of p electrons.

The Role of the Liquid–Liquid Interface in the Synthesis of Nonequilibrium Crystalline Wurtzite ZnS at Room Temperature

In this research, the role that the organic–inorganic liquid interface plays in the synthesis of nonequilibrium crystalline materials is investigated. A hierarchical nanocrystalline film of wurtzite ZnS, the high-temperature stable phase, is successfully prepared at room temperature by an interfacial in situ fabrication process. The organic–inorganic liquid interface constructed by n-hexane and water acts as the reaction zone for the synthesis of ZnS nanocrystalline film. A series of experimental results have proved that the liquid–liquid interface is the key factor for wurtzite ZnS formation at room temperature without any additive. The ZnS film consists of core–shell subunits characterized by ZnS nanoparticles around an organic core. Between the liquid–liquid interface, the core–shell subunits are coupled onto the surface of a SAM-modified substrate by terminal amino groups, so that the ZnS nanocrystalline film is formed by a layer-by-layer mode. This research brings forward a feasible route for synthesizing wurtzite ZnS in one-step process at room temperature and provides some beneficial information for studying the structural kinetics of nonequilibrium crystalline synthesis.

Comparative analysis of carboxysome shell proteins

Carboxysomes are metabolic modules for CO2 fixation that are found in all cyanobacteria and some chemoautotrophic bacteria. They comprise a semi-permeable proteinaceous shell that encapsulates ribulose-1,5-bisphosphate carboxylase/oxygenase (RuBisCO) and carbonic anhydrase. Structural studies are revealing the integral role of the shell protein paralogs to carboxysome form and function. The shell proteins are composed of two domain classes: those with the bacterial microcompartment (BMC; Pfam00936) domain, which oligomerize to form (pseudo)hexamers, and those with the CcmL/EutN (Pfam03319) domain which form pentamers in carboxysomes. These two shell protein types are proposed to be the basis for the carboxysome’s icosahedral geometry. The shell proteins are also thought to allow the flux of metabolites across the shell through the presence of the small pore formed by their hexameric/pentameric symmetry axes. In this review, we describe bioinformatic and structural analyses that highlight the important primary, tertiary, and quaternary structural features of these conserved shell subunits. In the future, further understanding of these molecular building blocks may provide the basis for enhancing CO2 fixation in other organisms or creating novel biological nanostructures.

Electronic structures and nonlinear optical properties of highly deformed halofullerenes C3v C60F18 and D3d C60Cl30

Electronic structures and nonlinear optical properties of two highly deformed halofullerenes C(3v) C(60)F(18) and D(3d) C(60)Cl(30) have been systematically studied by means of density functional theory. The large energy gaps (3.62 and 2.61 eV) between the highest occupied and lowest unoccupied molecular orbitals (HOMOs and LUMOs) and the strong aromatic character (with nucleus-independent chemical shifts varying from -15.08 to -23.71 ppm) of C(60)F(18) and C(60)Cl(30) indicate their high stabilities. Further investigations of electronic property show that C(60)F(18) and C(60)Cl(30) could be excellent electron acceptors for potential photonic/photovoltaic applications in consequence of their large vertical electron affinities. The density of states and frontier molecular orbitals are also calculated, which present that HOMOs and LUMOs are mainly distributed in the tortoise shell subunit of C(60)F(18) and the aromatic [18] trannulene ring of C(60)Cl(30), and the influence from halogen atoms is secondary. In addition, the static linear polarizability <alpha> and second-order hyperpolarizability <gamma> of C(60)F(18) and C(60)Cl(30) are calculated using finite-field approach. The values of <alpha> and <gamma> for C(60)F(18) and C(60)Cl(30) molecules are significantly larger than those of C(60) because of their lower symmetric structures and high delocalization of pi electrons.

P22 Coat Protein Structures Reveal a Novel Mechanism for Capsid Maturation: Stability without Auxiliary Proteins or Chemical Crosslinks

Viral capsid assembly and stability in tailed, dsDNA phage and Herpesviridae are achieved by various means including chemical crosslinks (unique to HK97), or auxiliary proteins (lambda, T4, phi29, and herpesviruses). All these viruses have coat proteins (CP) with a conserved, HK97-like core structure. We used a combination of trypsin digestion, gold labeling, cryo-electron microscopy, 3D image reconstruction, and comparative modeling to derive two independent, pseudoatomic models of bacteriophage P22 CP: before and after maturation. P22 capsid stabilization results from intersubunit interactions among N-terminal helices and an extensive "P loop," which obviate the need for crosslinks or auxiliary proteins. P22 CP also has a telokin-like Ig domain that likely stabilizes the monomer fold so that assembly may proceed via individual subunit addition rather than via preformed capsomers as occurs in HK97. Hence, the P22 CP structure may be a paradigm for understanding how monomers assemble in viruses like phi29 and HSV-1.

The Carboxysome and Other Bacterial Microcompartments

- Carboxysomes are part of the carbon concentrating mechanism in cyanobacteria and chemoautotrophs. - Carboxysomes are a subclass of bacterial microcompartments (BMCs); BMCs can encapsulate a range of metabolic processes. - Like some viral particles, the carboxysome can be modeled as an icosahedron-in its case, having 4,000-5,000 hexameric shell subunits and 12 surface pentamers to generate curvature. - The threefold axis of symmetry of the CsoS1D protein in carboxysomes forms a pore that can open and close, allowing for selective diffusion. - Genetic modules encoding BMC shell proteins and the enzymes that they encapsulate are horizontally transferable, suggesting they enable bacteria to adapt to diverse environments.

Behavioral inhibition during reversal learning in the limbic system: a computational model

A biological model of the sub-cortical nuclei of the limbic system capable of performing reversal learning in a foodseeking task is presented. Contrary to other learning models, the reversal is modeled not by eliminating (unlearning) learned behaviors, but rather by the inhibiting the previously learned behavior. This allows for the re-instatement of behavior to be quickly re-established in such a way that animal behavior is simulated. In this model, the role of dopamine is different to standard models. The activity of dopaminergic (DA) neurons required for successful conditioning has commonly been identified as an error signal whereby an increased DA activity codes a positive error i.e. long term potentiation (LTP) and a decrease in dopamine concentration codes a negative error. In this model learning is achieved by implementing a form of differential Hebbian learning known as Isotropic Sequence Order learning and a third factor (ISO3). This third factor enables learning to be triggered at relevant moments. It is modeled by dopaminergic neurons which can be activated via a direct excitatory glutamatergic pathway, and an indirect disinhibitory GABAergic pathway. While the former generates phasic DA release which during acquisition enables long term potentiation (LTP) to occur, the latter produces in an increased population of tonically active DA neurons which generates long term depression (LTD) when an adjustment in learned behavior is required. The nucleus accumbens is divided into two shell and core subunits and has been modeled to function in distinct but complementary manners. Here, the core uses conditioned reinforcers to invigorate instrumental responding. The core has been modeled to learn to enable behavioral responding to reward predicting stimuli and as such undergoes minimal LTD so as not to eliminate these learned processes. On the other hand, LTD occurs in the shell that, through a shell-ventral pallido-medio dorsal pathway, influences the core and enables behavioral flexibility. A simple reversal-learning task will be used to demonstrate how this biophysically realistic pathway can be used to learn and reverse a simple food-seeking task. from Eighteenth Annual Computational Neuroscience Meeting: CNS*2009 Berlin, Germany. 18–23 July 2009

Structural transitions in the scaffolding and coat proteins of P22 virus during assembly and disassembly.

An in vitro system for investigating the assembly of the Salmonella phage P22 has been exploited to elucidate the structural basis of recognition between scaffolding protein (gp8) and coat protein (gp5) subunits of the viral procapsid. Raman spectroscopy and circular dichroism have been employed to examine structural thermostabilities of both gp8 and gp5 in native procapsids, and to characterize structural changes accompanying scaffolding exit, procapsid expansion, and shell disassembly. It is found that the secondary structure of the isolated gp8 subunit is rich in alpha-helix (approximately 40%), is highly thermolabile, and is characterized by noncooperative unfolding (Tm approximately 49 degrees C). Conversely, the procapsid-bound gp8 subunit exhibits stabilization of its alpha-helical secondary structure, characterized by cooperative unfolding. Because cooperative unfolding of gp8 coincides with exit from the procapsid, the present results suggest that unfolding and release are coupled processes. Structural differences between procapsid-free and procapsid-bound gp8 subunits are also apparent in Raman markers which monitor environments of tyrosine and tryptophan side chains. Temperature-resolved Raman spectroscopy of the empty procapsid shell reveals three distinct structural transitions for the gp5 subunits. The first, which occurs between 50 and 65 degrees C, is attributed to shell expansion and results in an increase in beta-strand secondary structure. The two higher temperature transitions, occurring within intervals of 70-80 and 80-95 degrees C, respectively, are attributed to partial unfolding of the shell subunit and subsequent shell disassembly. The same gp5 structure transitions are detected for procapsids which contain scaffolding protein. On the basis of the observed thermodynamic coupling between gp8 unfolding and its release from the procapsid, we propose a model for P22 procapsid assembly. Implications of the model for in vivo assembly of dsDNA viruses are discussed.