Structural and biochemical characterization of a widespread enterobacterial peroxidase encapsulin.

Encapsulins are self-assembling protein compartments found in prokaryotes and specifically encapsulate dedicated cargo enzymes. The most abundant encapsulin cargo class are Dye-decolorizing Peroxidases (DyPs). It has been previously suggested that DyP encapsulins are involved in oxidative stress resistance and bacterial pathogenicity due to DyPs' inherent ability to reduce and detoxify hydrogen peroxide while oxidizing a broad range of organic co-substrates.

The biosynthesis of the odorant 2-methylisoborneol is compartmentalized inside a protein shell.

Terpenoids are the largest class of natural products, found across all domains of life. One of the most abundant bacterial terpenoids is the volatile odorant 2-methylisoborneol (2-MIB), partially responsible for the earthy smell of soil and musty taste of contaminated water. Many bacterial 2-MIB biosynthetic gene clusters were thought to encode a conserved transcription factor, named EshA in the model soil bacterium Streptomyces griseus.

Pore Engineering as a General Strategy to Improve Protein-Based Enzyme Nanoreactor Performance.

Enzyme nanoreactors are nanoscale compartments consisting of encapsulated enzymes and a selectively permeable barrier. Sequestration and colocalization of enzymes can increase catalytic activity, stability, and longevity, highly desirable features for many biotechnological and biomedical applications of enzyme catalysts. One promising strategy to construct enzyme nanoreactors is to repurpose protein nanocages found in nature.

Pore engineering as a general strategy to improve protein-based enzyme nanoreactor performance.

Enzyme nanoreactors are nanoscale compartments consisting of encapsulated enzymes and a selectively permeable barrier. Sequestration and co-localization of enzymes can increase catalytic activity, stability, and longevity, highly desirable features for many biotechnological and biomedical applications of enzyme catalysts. One promising strategy to construct enzyme nanoreactors is to repurpose protein nanocages found in nature.

Dissecting the stability of Atezolizumab with renewable amino acid-based ionic liquids: Colloidal stability and anticancer activity under thermal stress.

Monoclonal antibodies (mAbs) have revolutionised the biopharmaceutical market. Being proteinaceous, mAbs are prone to chemical and physical instabilities. Various approaches were attempted to stabilise proteins against degradation factors.

A two-component quasi-icosahedral protein nanocompartment with variable shell composition and irregular tiling.

Protein shells or capsids are a widespread form of compartmentalization in nature. Viruses use protein capsids to protect and transport their genomes while many cellular organisms use protein shells for varied metabolic purposes. These protein-based compartments often exhibit icosahedral symmetry and consist of a small number of structural components with defined roles.

Cyclodipeptide oxidase is an enzyme filament.

Modified cyclic dipeptides represent a widespread class of secondary metabolites with diverse pharmacological activities, including antibacterial, antifungal, and antitumor. Here, we report the structural characterization of the Streptomyces noursei enzyme AlbAB, a cyclodipeptide oxidase (CDO) carrying out α,β-dehydrogenations during the biosynthesis of the antibiotic albonoursin. We show that AlbAB is a megadalton heterooligomeric enzyme filament containing covalently bound flavin mononucleotide cofactors.

Structural basis for peroxidase encapsulation inside the encapsulin from the Gram-negative pathogen Klebsiella pneumoniae.

Encapsulins are self-assembling protein nanocompartments capable of selectively encapsulating dedicated cargo proteins, including enzymes involved in iron storage, sulfur metabolism, and stress resistance. They represent a unique compartmentalization strategy used by many pathogens to facilitate specialized metabolic capabilities. Encapsulation is mediated by specific cargo protein motifs known as targeting peptides (TPs), though the structural basis for encapsulation of the largest encapsulin cargo class, dye-decolorizing peroxidases (DyPs), is currently unknown.

Point mutation in a virus-like capsid drives symmetry reduction to form tetrahedral cages.

Protein capsids are a widespread form of compartmentalisation in nature. Icosahedral symmetry is ubiquitous in capsids derived from spherical viruses, as this geometry maximises the internal volume that can be enclosed within. Despite the strong preference for icosahedral symmetry, we show that simple point mutations in a virus-like capsid can drive the assembly of novel symmetry-reduced structures.

A widespread bacterial protein compartment sequesters and stores elemental sulfur.

Subcellular compartments often serve to store nutrients or sequester labile or toxic compounds. As bacteria mostly do not possess membrane-bound organelles, they often have to rely on protein-based compartments. Encapsulins are one of the most prevalent protein-based compartmentalization strategies found in prokaryotes.

Cyclodipeptide oxidase is an enzyme filament.

Modified cyclic dipeptides represent a widespread class of secondary metabolites with diverse pharmacological activities, including antibacterial, antifungal, and antitumor. Here, we report the structural characterization of the enzyme AlbAB, a cyclodipeptide oxidase (CDO) carrying out α,β-dehydrogenations during the biosynthesis of the antibiotic albonoursin. We show that AlbAB is a megadalton heterooligomeric enzyme filament containing covalently bound flavin mononucleotide cofactors.

Structural basis for peroxidase encapsulation in a protein nanocompartment.

Encapsulins are self-assembling protein nanocompartments capable of selectively encapsulating dedicated cargo proteins, including enzymes involved in iron storage, sulfur metabolism, and stress resistance. They represent a unique compartmentalization strategy used by many pathogens to facilitate specialized metabolic capabilities. Encapsulation is mediated by specific cargo protein motifs known as targeting peptides (TPs), though the structural basis for encapsulation of the largest encapsulin cargo class, dye-decolorizing peroxidases (DyPs), is currently unknown.

HIV-1 binds dynein directly to hijack microtubule transport machinery.

Viruses exploit host cytoskeletal elements and motor proteins for trafficking through the dense cytoplasm. Yet the molecular mechanism that describes how viruses connect to the motor machinery is unknown. Here, we demonstrate the first example of viral microtubule trafficking from purified components: HIV-1 hijacking microtubule transport machinery.

Structure and heterogeneity of a highly cargo-loaded encapsulin shell.

Encapsulins are self-assembling protein nanocompartments able to selectively encapsulate dedicated cargo enzymes. Encapsulins are widespread across bacterial and archaeal phyla and are involved in oxidative stress resistance, iron storage, and sulfur metabolism. Encapsulin shells exhibit icosahedral geometry and consist of 60, 180, or 240 identical protein subunits.

Structure and heterogeneity of a highly cargo-loaded encapsulin shell.

Encapsulins are self-assembling protein nanocompartments able to selectively encapsulate dedicated cargo enzymes. Encapsulins are widespread across bacterial and archaeal phyla and are involved in oxidative stress resistance, iron storage, and sulfur metabolism. Encapsulin shells exhibit icosahedral geometry and consist of 60, 180, or 240 identical protein subunits.

Exploring the Extreme Acid Tolerance of a Dynamic Protein Nanocage.

Encapsulins are microbial protein nanocages capable of efficient self-assembly and cargo enzyme encapsulation. Due to their favorable properties, including high thermostability, protease resistance, and robust heterologous expression, encapsulins have become popular bioengineering tools for applications in medicine, catalysis, and nanotechnology. Resistance against physicochemical extremes like high temperature and low pH is a highly desirable feature for many biotechnological applications.

Warmth and competence perceptions of key protagonists are associated with containment measures during the COVID-19 pandemic: Evidence from 35 countries.

It is crucial to understand why people comply with measures to contain viruses and their effects during pandemics. We provide evidence from 35 countries (N = 12,553) from 6 continents during the COVID-19 pandemic (between 2021 and 2022) obtained via cross-sectional surveys that the social perception of key protagonists on two basic dimensions-warmth and competence-plays a crucial role in shaping pandemic-related behaviors. Firstly, when asked in an open question format, heads of state, physicians, and protest movements were universally identified as key protagonists across countries.

Heterologous expression and purification of encapsulins in .

In recent years a large number of encapsulin nanocompartment-encoding operons have been identified in bacterial and archaeal genomes. Encapsulin-encoding genes and operons from GC-rich Gram-positive bacteria, particularly of the phylum Actinobacteria, are often difficult to overexpress and purify in a soluble form using standard expression systems Here, we present a protocol to heterologously overexpress encapsulin nanocompartments and encapsulin-containing operons in Successful encapsulin production begins with the transfer of a expression plasmid, encoding the gene(s) of interest, via conjugation to the model actinobacterium After growing the conjugated culture to the optimal optical density, protein production is induced by the addition of the inducer thiostrepton, followed by expression in liquid culture for 1-3 days. Cells are lysed and encapsulin proteins purified using ammonium sulfate precipitation and size exclusion chromatography.

Synthesis and Reconstitution Using Mammalian Cell-Free Lysates Enables the Systematic Study of the Regulation of LINC Complex Assembly.

Understanding the structure and structure-function relationships of membrane proteins is a fundamental problem in biomedical research. Given the difficulties inherent to performing mechanistic biochemical and biophysical studies of membrane proteins , we previously developed a facile HeLa cell-based cell-free expression (CFE) system that enables the efficient reconstitution of full-length (FL) functional inner nuclear membrane Sad1/UNC-84 (SUN) proteins (i.e.

Pore structure controls stability and molecular flux in engineered protein cages.

Protein cages are a common architectural motif used by living organisms to compartmentalize and control biochemical reactions. While engineered protein cages have featured in the construction of nanoreactors and synthetic organelles, relatively little is known about the underlying molecular parameters that govern stability and flux through their pores. In this work, we systematically designed 24 variants of the encapsulin cage, featuring pores of different sizes and charges.

Triggered Reversible Disassembly of an Engineered Protein Nanocage*.

Protein nanocages play crucial roles in sub-cellular compartmentalization and spatial control in all domains of life and have been used as biomolecular tools for applications in biocatalysis, drug delivery, and bionanotechnology. The ability to control their assembly state under physiological conditions would further expand their practical utility. To gain such control, we introduced a peptide capable of triggering conformational change at a key structural position in the largest known encapsulin nanocompartment.

Large-scale computational discovery and analysis of virus-derived microbial nanocompartments.

Encapsulins are a class of microbial protein compartments defined by the viral HK97-fold of their capsid protein, self-assembly into icosahedral shells, and dedicated cargo loading mechanism for sequestering specific enzymes. Encapsulins are often misannotated and traditional sequence-based searches yield many false positive hits in the form of phage capsids. Here, we develop an integrated search strategy to carry out a large-scale computational analysis of prokaryotic genomes with the goal of discovering an exhaustive and curated set of all HK97-fold encapsulin-like systems.