Yutetsu Kuruma

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Cell-free translation systems have developed significantly over the last two decades and improvements in yield have resulted in their use for protein production in the laboratory. These systems have protein engineering applications, such as the production of proteins containing unnatural amino acids and development of proteins exhibiting novel functions.(More)
In the field of molecular biology or biochemistry, preparation and use of purified proteins involved in a certain biological system is crucial for understanding their mechanisms and functions in cells or organisms. The recent progress in a cell-free translation system allows us to prepare proteins in a test tube directly from cDNAs that encode the amino(More)
Synthetic biology is an emerging field that aims at constructing artificial biological systems by combining engineering and molecular biology approaches. One of the most ambitious research line concerns the so-called semi-synthetic minimal cells, which are liposome-based system capable of synthesizing the lipids within the liposome surface. This goal can be(More)
By combining translation and membrane integration/translocation systems, we have constructed a novel cell-free system for the production of presecretory and integral membrane proteins in vitro. A totally defined, cell-free system reconstituted from a minimal number of translation factors was supplemented with urea-washed inverted membrane vesicles (U-INVs)(More)
The a subunit of F(1)F(o) (F(1)F(o)-ATP synthase) is a highly hydrophobic protein with five putative transmembrane helices which plays a central role in H(+)-translocation coupled with ATP synthesis/hydrolysis. In the present paper, we show that the a subunit produced by the in vitro protease-free protein synthesis system (the PURE system) is integrated(More)
In a significant step towards the construction of the semi-synthetic minimal cell, a protein expression system with a minimal set of pure and specific enzymes is required. A novel cell-free transcription and translation system named PURESYSTEM (PS), consisting of a specified set of 36 enzymes and ribosomes, has been entrapped in POPC liposomes for protein(More)
In F(o)F(1)-ATP synthase, multimeric c-subunits are assembled to a ring (c-ring) in the membranes that rotates as protons flow across F(o). We recently reported that assembly of c-ring of Propionigenium modestum in the membranes of Escherichia coli cells required P. modestum UncI, a product of the conserved uncI gene in the F(o)F(1) operon. However,(More)
Self-reproduction is one of main properties that define living cells. In order to explore the self-reproduction process for the study of early cells, and to develop a research line somehow connected to the origin of life, we have built up a constructive ‘synthetic cells (minimal cells)’ approach. The minimal cells approach consists in the investigation of(More)
Protein expression is the most complex metabolic reaction that has been encapsulated in liposomes, mainly as an intermediate step toward the synthesis of minimal semisynthetic cells. Although there are different experimental approaches to achieving the synthesis of proteins inside liposomes and it is therefore not possible to give a standard recipe, all(More)
The compartmentalization of a cell-free gene expression system inside a self-assembled lipid vesicle is envisioned as the simplest chassis for the construction of a minimal cell. Although crucial for its realization, quantitative understanding of the dynamics of gene expression in bulk and liposome-confined reactions is scarce. Here, we used two orthogonal(More)