The Potential of Artificial Cells Functioning under Deep-Sea Conditions.

Artificial cells with reconstructed cellular functions could serve as practical protocell models for studying the early cellular life on the Earth. Investigating the viability of protocell models in extreme environments where life may have arisen is important for advancing origin-of-life research. Here, we tested the survivability of lipid membrane vesicles in deep-sea environments.

Regulated N-Terminal Modification of Proteins Synthesized Using a Reconstituted Cell-Free Protein Synthesis System.

The N-terminal modification of nascent proteins, such as acetylation and myristoylation, is one of the most abundant post-translational modifications. To analyze the function of the modification, it is important to compare the modified and unmodified proteins under defined conditions. However, it is technically difficult to prepare unmodified proteins because cell-based systems contain endogenous modification systems.

Phospholipid synthesis inside phospholipid membrane vesicles.

Construction of living artificial cells from genes and molecules can expand our understanding of life system and establish a new aspect of bioengineering. However, growth and division of cell membrane that are basis of cell proliferation are still difficult to reconstruct because a high-yielding phospholipid synthesis system has not been established. Here, we developed a cell-free phospholipid synthesis system that combines fatty acid synthesis and cell-free gene expression system synthesizing acyltransferases.

Proteomic Time-Course Analysis of the Filamentous Anoxygenic Phototrophic Bacterium, , during the Transition from Respiration to Phototrophy.

is a filamentous anoxygenic phototrophic bacterium that grows chemotrophically under oxic conditions and phototrophically under anoxic conditions. Because photosynthesis-related genes are scattered without any gene clusters in the genome, it is still unclear how this bacterium regulates protein expression in response to environmental changes. In this study, we performed a proteomic time-course analysis of how expresses proteins to acclimate to environmental changes, namely the transition from chemoheterotrophic respiratory to photoheterotrophic growth mode.

Rapid and Facile Preparation of Giant Vesicles by the Droplet Transfer Method for Artificial Cell Construction.

Giant vesicles have been widely used for the bottom-up construction of artificial (or synthetic) cells and the physicochemical analysis of lipid membranes. Although methods for the formation of giant vesicles and the encapsulation of molecules within them have been established, a standardized protocol has not been shared among researchers including non-experts. Here we proposed a rapid and facile protocol that allows the formation of giant vesicles within 30 min.

Phylogenomics of Haloarchaea: The Controversy of the Genera .

The haloarchaeal genera and were described almost simultaneously by two different research groups and some strains studied separately were described as different species of these genera. Furthermore, the description of additional species were assigned to either or , mainly on the basis of the phylogenetic comparative analysis of single genes (16S rRNA gene and more recently gene), but these species were not adequately separated or assigned to the corresponding genus. Some studies suggested that the species of these two genera should be unified into a single genus, while other studies indicated that the genera should remain but some of the species should be reassigned.

sp. nov., a galactomannan-degrading haloarchaeon isolated from commercial salt.

A mannan-degrading halophilic archaeal strain, MD130-1, was isolated from a commercial salt sample. Cells were motile, rod-shaped, and stained Gram-negative. Colonies were pink pigmented.

Structural comparisons of phosphoenolpyruvate carboxykinases reveal the evolutionary trajectories of these phosphodiester energy conversion enzymes.

Inorganic pyrophosphate (PP) consists of two phosphate molecules and can act as an energy and phosphate donor in cellular reactions, similar to ATP. Several kinases use PP as a substrate, and these kinases have recently been suggested to have evolved from ATP-dependent functional homologs, which have significant amino acid sequence similarity to PP-utilizing enzymes. In contrast, phosphoenolpyruvate carboxykinase (PEPCK) can be divided into three types according to the phosphate donor (ATP, GTP, or PPi), and the amino acid sequence similarity of these PEPCKs is too low to confirm that they share a common ancestor.

Accurate high-throughput screening based on digital protein synthesis in a massively parallel femtoliter droplet array.

We report a general strategy based on digital counting principle that enables an efficient acquisition of enzyme mutants with desired activities from just a few clones within a day. We prepared a high-density femtoliter droplet array, consisting of 1 million uniform droplets per 1 cm to carry out high-throughput protein synthesis and screening. Single DNA molecules were randomly distributed into each droplet following a Poisson process to initiate the protein synthesis with coupled cell-free transcription and translation reactions and then recovered by a microcapillary.

Computational Analysis of the Interaction Energies between Amino Acid Residues of the Measles Virus Hemagglutinin and Its Receptors.

Measles virus (MV) causes an acute and highly devastating contagious disease in humans. Employing the crystal structures of three human receptors, signaling lymphocyte-activation molecule (SLAM), CD46, and Nectin-4, in complex with the measles virus hemagglutinin (MVH), we elucidated computationally the details of binding energies between the amino acid residues of MVH and those of the receptors with an fragment molecular orbital (FMO) method. The calculated inter-fragment interaction energies (IFIEs) revealed a number of significantly interacting amino acid residues of MVH that played essential roles in binding to the receptors.

Complete Genome Sequence of Sphingobium sp. Strain YG1, a Lignin Model Dimer-Metabolizing Bacterium Isolated from Sediment in Kagoshima Bay, Japan.

sp. strain YG1 is a lignin model dimer-metabolizing bacterium newly isolated from sediment in Kagoshima, Japan, at a depth of 102 m. Here, we report the complete genome nucleotide sequence of strain YG1.

Salinarchaeum chitinilyticum sp. nov., a chitin-degrading haloarchaeon isolated from commercial salt.

Two chitin-degrading halophilic archaeal strains, MC-74T and MC-23, were isolated from commercial salt samples. Cells were motile, rod-shaped and stained Gram-negative. Colonies were vermillion-pigmented.

Haloparvum alkalitolerans sp. nov., alkali-tolerant haloarchaeon isolated from commercial salt.

A Gram-stain-negative, rod-pleomorphic, aerobic, halophilic archaeon, strain MK62-1T, was isolated from commercial salt made from seawater in the Philippines. Strain MK62-1T was able to grow at 2.1-4.

Halopiger thermotolerans sp. nov., a thermo-tolerant haloarchaeon isolated from commercial salt.

Three thermo-tolerant halophilic archaeal strains, SR-441T, SR-412 and SR-188, were isolated from commercial salt samples. Cells were non-motile pleomorphic rod-shaped, and stained Gram-negative. Colonies were pink-pigmented.

Halarchaeum grantii sp. nov., a moderately acidophilic haloarchaeon isolated from a commercial salt sample.

Three moderately acidophilic, halophilic archaeal strains, MH1-243-3T, MH1-243-5 and MH1-243-6, were isolated from a commercial salt sample made from seawater in Okinawa, Japan. Cells of the three strains were pleomorphic and stained Gram-negative. Colonies of the strains were orange-red-pigmented.

Halorubrum gandharaense sp. nov., an alkaliphilic haloarchaeon from commercial rock salt.

A Gram-stain-negative, non-motile, pleomorphic rod-shaped, orange-red-pigmented, facultatively aerobic and haloalkaliphilic archaeon, strain MK13-1T, was isolated from commercial rock salt imported from Pakistan. The NaCl, pH and temperature ranges for growth of strain MK13-1T were 3.0-5.

Aneurinibacillus tyrosinisolvens sp. nov., a tyrosine-dissolving bacterium isolated from organics- and methane-rich seafloor sediment.

A novel Gram-positive-staining, strictly aerobic and heterotrophic bacterium, designated strain LL-002T, was isolated from organics- and methane-rich seafloor sediment at a depth of 100 m in Kagoshima Bay, Kagoshima, Japan. Colonies were lustreless and translucent white in colour. The temperature, pH and salt concentration ranges for growth were 10-30 °C, pH 6.

Halocalculus aciditolerans gen. nov., sp. nov., an acid-tolerant haloarchaeon isolated from commercial salt.

Three halophilic archaeal strains, MH2-243-1(T), MH2-93-1 and MH2-91-1 were isolated from commercial salt samples from Japan, Australia, and Bolivia. Strain MH2-243-1(T) was able to grow in the presence of 12-30% (w/v) NaCl (optimum, 18% NaCl), at pH 4.5-7.

Halococcus agarilyticus sp. nov., an agar-degrading haloarchaeon isolated from commercial salt.

Two agar-degrading halophilic archaeal strains, 62 E(T) and 197 A, were isolated from commercial salt samples. Cells were non-motile cocci, approximately 1.2-2.

Structurally distinct hybrid polymer/lipid nanoconstructs harboring a type-I ribotoxin as cellular imaging and glioblastoma-directed therapeutic vectors.

A nanoformulation composed of a ribosome inactivating protein-curcin and a hybrid solid lipid nanovector has been devised against glioblastoma. The structurally distinct nanoparticles were highly compatible to human endothelial and neuronal cells. A sturdy drug release from the particles, recorded upto 72 h, was reflected in the time-dependent toxicity.

Alicyclobacillus cellulosilyticus sp. nov., a thermophilic, cellulolytic bacterium isolated from steamed Japanese cedar chips from a lumbermill.

A thermophilic bacterium, strain Sueoka(T), was isolated from steamed Japanese cedar chips from a lumber mill in Gobo, Japan. The strain was able to grow on carboxymethyl cellulose at 60 °C, was Gram-stain-negative, and grew between 40.0 and 67.

Enhancing thermostability and the structural characterization of Microbacterium saccharophilum K-1 β-fructofuranosidase.

A β-fructofuranosidase from Microbacterium saccharophilum K-1 (formerly known as Arthrobacter sp. K-1) is useful for producing the sweetener lactosucrose (4(G)-β-D-galactosylsucrose). Thermostability of the β-fructofuranosidase was enhanced by random mutagenesis and saturation mutagenesis.