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Developing Volume-CAD as the basis for next generation manufacturing
RIKEN Center for Intellectual Property Programs and Management |
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| What is CAD? | |||
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Kiwamu Kase, laboratory head of the Volume-CAD Development Team, explains: "CAD is an abbreviation for computer-aided design, and refers to the process of designing products using a computer." In the past, design drawings were always prepared by hand. Because of this, conventional drawings only show sectional views, but all the necessary data to build the product are contained in the drawings. The craftsman imagines the shape of the finished product from the drawings and works to create it. This requires skill, and unfortunately in recent times Japan is suffering from a lack of young people who are trained as craftsmen. "What should we do to free ourselves from relying on craftsmanship? The answer is to replace all the information given in the drawings with data that can be computer-processed, and to represent the product in three-dimensions, rather than as sectional views, in a way that will enable anyone to picture what the product will look like. This is the essence of CAD." The concept of CAD emerged in the 1960's. Since then, CAD has seen constant increases in the precision with which we can represent complex three-dimensional shapes (Figure 1). Three models have been developed to date. The earliest is the wire frame model, which represents shapes with curved lines drawn by joining points in three-dimensional space. The next to be developed was the surface model, a development from the wire frame model, with a skin only covering a portion of its surface. The most advanced is the solid model, which has a skin covering the entire surface to enable differentiation between the inside and the outside. CAD is now essential to the manufacturing process, and is used to design a broad range of products for society, including automobiles, equipment, and buildings.
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| Integrating information on making products | |||
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Today, design is not the only process involved in making products using computers. Making a product involves design, analysis, manufacturing, and testing, and in each of these processes computers are used. These computer-aided processes are called computer-aided design (CAD), computer-aided engineering (CAE), computer-aided manufacturing (CAM), and computer-aided testing (CAT), respectively. Because of the different software programs used in these individual processes, however, it has become impossible to handle integrated data. Losing data due to errors during data conversion is also a concern. The greatest problem with conventional CAD resides in its inability to achieve appropriate simulations. When making a product, it is necessary to carry out various experiments to analyze its performance, toughness, and other features. For this purpose, the experimental work requires a prototype. In addition, feedback cycles must be repeated to make corrections in the initial design based on experimental results, and to test new trial products. This process takes a great deal of time and money. The problem can be resolved by various simulations. Simulating experiments on a computer, without making a prototype, would dramatically reduce time and costs. Kase points out, however, that "conventional surface or solid CAD data cannot be used directly for simulation. Although the solid model seems to reflect reality accurately, it is quite different from the actual product, which is formed from different materials such as metal and plastic, each with its own properties. However, the solid model comprises a skin only and does not represent the material content, despite its identity as a tool for three-dimensional CAD." Although a solid model may be used for simulation, constructing one reportedly needs a lot of very detailed work, including dividing the entire shape into fine elements known as meshes. In addition, the complicated procedure of adding and removing parts can result in an accumulation of numerical errors that can result in a hole and hence the model fails. "With this in mind, I proposed developing true three-dimensional CAD with greater realism, that models the whole material content, and which can be used directly in simulations, and that was the beginning of Volume-CAD (VCAD)." Following on from this, RIKEN started its Integrated Volume-CAD System Research Program (Program Director: Akitake Makinouchi) in April 2001. The program has six teams: Volume-CAD Development, V-CAT Development, Product Performance Simulation, Manufacturing Process Simulation, V-CAD Fabrication, and V-CAD Technology Transfer. "As seen from the existence of the V-CAD Technology Transfer Team, one task of the project is to find practical applications for its achievements, as well as to do research and present papers." Entrusted with the project's core task of developing Volume-CAD, team leader Kase took up the challenge, with the idea of establishing an innovative system. "I set as my goal the development of a new system that should serve as the basis for next generation manufacturing, and that would be a complete integration of CAD, simulations, CAM, and CAT." |
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| "Kitta Cube" -- cut cube | |||
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"Our starting point for Volume-CAD lies in the idea of filling the model with voxel cubes," says Kase. The term "pixel" refers to the basic unit used to represent two-dimensional images, and the "voxel" is the unit used to represent three-dimensional images. Because a solid model is a continuous body, it is impossible to extract a portion of it. On the other hand, the voxel model has the advantage that it permits separate processing of discrete cubes using more than one computer because the individual cubes can be numbered. When handling large amounts of data, parallel processing is important. However, since it consists simply of stacked cubes, the conventional voxel model is unattractive to look at because of its irregular surfaces (Figure 1). To overcome this drawback, Kase and team member Yoshinori Teshima jointly devised a new approach. "To obtain smooth surfaces, an oblique plane is necessary after all. So, we enclosed a triangular plane in the cube." They assigned one (cutting) point to each line (edge) of the cube and joined three of the points to form a triangle that partitions the cube (Figure 2). This approach is drastic as it approximates complicated shapes with triangles. Teshima has demonstrated that 23,520 patterns are available in terms of the positioning of the cutting points, and that as many as 1064 cube partitioning patterns are possible. As is evident from Figure 1, the volume model is capable of representing finer shapes than the voxel model. Kase and Teshima called this method the "Kitta Cube." "Kitta" is a Japanese adjective that means "cut," and so the term "Kitta Cube" means a cut cube. In the hope of signifying worldwide that the new idea originated in Japan, they combined the Japanese and English words to make a new term.
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| Cell hierarchization | |||
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"The most important feature of Volume-CAD is the inclusion of not only shape information but also physical attribute data in the cubes, which are called cells," says Kase. Physical attributes refer to various variables, such as hardness, temperature, and strain. Conventional CAD is intended to represent shapes, not types incorporating physical attributes. "Representing shapes and physical attributes within the same framework seems to be an idea that everyone can appreciate easily but has not yet been achieved." There are two types of Volume-CAD cells: those with a boundary (boundary cells) and those without (non-boundary cells) (Figure 3). Because a boundary is also an outline of an object, the boundary cell carries shape information and two or more sets, outer and inner, and so on, of physical attributes; e.g., air for the outside and iron for the inside. On the other hand, the non-boundary cell carries only one set of physical attributes. Stacking boundary and non-boundary cells enables us to represent multi-component media such as aluminum in iron. In the second year of the project, Kase carried out a major update of the software program then under development. "A hierarchical version of the program, allowing cell size changes according to position, was developed under the lead of Shugo Usami, a visiting team member from a commercial company. With greater differentials of shapes and physical attributes, finer cells are easier to handle. We have made it possible (Figure 4)." Although fine shapes are better represented by stacking smaller cells of the same size, that approach is wasteful due to the unwanted distribution of such cells in spaces where nothing is otherwise present. Data volume can be reduced using the octree method, which finely divides only those portions undergoing frequent morphological changes or major changes in physical properties. As each side of the cell is halved, the cell is divided into eight segments. This is the principle of the octree method. "Two big features of Volume-CAD reside in the incorporation of planes in each cell and the hierarchization of the cells. No other type of CAD incorporates this idea. We wanted to make a new system from our own ideas." In the third year, they constructed a framework for Volume-CAD. "The framework can be compared to an infrastructure like a road. We intend to provide a function that offers free access to anyone who wants to use it." Currently, an investigation to establish a system for parallel data processing using several computers is ongoing under the leadership of team member Masaya Kato.
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| Potential for simulations of casting and injection molding | |||
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To date, we have filed more than 30 patent applications concerning Volume-CAD from the Integrated V-CAD System Research Program. Some patented systems are already in use by commercial companies, including several products generally available on the market. We release development versions of our Volume-CAD software program to other teams. In addition, we have recently established a "Volume-CAD System Study Association" and encourage its members (enterprises and scientists) to use the program, and to report back their findings to enable us to improve our developmental activities. "Many members of the Association realize the potential of Volume-CAD for casting and plastic injection molding and want to use the software," Kase says in confidence. In manufacturing, it is common practice in casting and injection molding to design and make a metal mold, pour molten metal or plastic material into the mold and allow it to solidify. However, the actual molding operation sometimes results in deformation or cracking of the mold, or cavitation at important places. In all these cases, the mold must be repaired. Using Volume-CAD, it is possible to optimize the mold shape by means of a simulation, without actually pouring the molten metal into the mold. Because Volume-CAD enables us to simulate the flow of metal during casting, as well as the finished shape, it is easier to identify the causes of molding failures and to come up with solutions. Simulations with Volume-CAD have been found to reflect experimental results better than those performed using conventional meshes with rectangular lattices (Figure 5). This is a good example of one of the remarkable features of Volume-CAD, i.e., that it is capable of naturalistically representing both flow paths of objects with unclear boundaries like water and oblique flow paths with desirably clear boundaries.
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| Expanding the applicability of Volume-CAD | |||
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Although Volume-CAD is being developed with the aim of establishing a basic system for next generation manufacturing, its targets are not limited to man-made products. It will find new applications, even to diastrophism and human blood streams. Team member Yutaka Otake, who joined the team last year, is expected to take an active part in the plan. "Another strong point of Volume-CAD is its versatile applicability to both man-made and natural products, and even to both solid and liquid simulations." Finally, Kase talked about the future of Volume-CAD: "In the future, our Volume-CAD software program will be offered to the public for free. I am happy to see the program helping students and young scientists to create new products." 
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Determining protein structures using the electron microscope and contributing to medical practice
RIKEN Harima Institute |
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| Acetylcholine receptor | |||
 "Elucidation of the mechanism of life from protein functions is the ultimate goal of our work," Miyazawa began his talk. Then, what is life?"I think that the state of 'living' or 'being alive' can be defined as responding to external stimuli. These responses come from the actions of individual cells, which are based on protein functions. In exploring the mechanism of life, it is important to investigate the mechanism of protein functions." Miyazawa has been working to elucidate the nicotinic acetylcholine receptor, which plays a key role in responding to many stimuli. The acetylcholine receptor, embedded in the cell membrane of the muscle cell, receives acetylcholine released from the terminus of a nerve cell (Figure 1). As acetylcholine binds to the receptor, an ion channel opens, through which positively charged sodium ions (Na+) enter from outside the cell, resulting in electrical signals. Information from the nerve cells is thus transmitted to cause muscle contraction. Chemical substances that transmit nerve cell signals, like acetylcholine, are called "neurotransmitters." To date, several tens of kinds of neurotransmitters have been found, and acetylcholine was the first to be discovered in 1921. In 1970, the acetylcholine receptor was discovered, followed by the determination of its amino acid sequence (primary structure) in 1982 by Shosaku Numa, then professor at Kyoto University, and his colleagues. Essentially, a protein is composed of a chain of approximately 20 kinds of amino acid residues based on their genetic information. Prof. Numa and his colleagues elucidated the arrangement of amino acids of acetylcholine receptor. However, the primary structure was insufficient to elucidate the functional mechanism of the acetylcholine receptors, because no protein can function normally unless it forms a three-dimensional structure which is the result of the complex folding of one-dimensional sequences of amino acids. |
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| Elucidation of the structure using electron microscope | |||
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In 1984, Dr. Nigel Unwin and colleagues of the Laboratory of Molecular Biology, Medical Research Council (MRC) in the UK began work to determine the structure of the acetylcholine receptor. In 1995, Miyazawa joined their group. He was subsequently able to determine the structure in full cooperation with Yoshinori Fujiyoshi (Professor at Kyoto University), Group Director of the Membrane Dynamics Research Group of RIKEN Harima Institute, in 2003. It took them almost twenty years to determine the structure. What difficulties did they encounter? A representative approach to the structural analysis of protein is X-ray crystallography. In this method, a protein crystal is exposed to X-rays, and its three-dimensional structure is calculated from diffraction images of a large number of protein molecules in the crystal. Crystal diffraction requires the subject protein to be crystallized in the first step. When the cell membrane, which contains the acetylcholine receptor, is crystallized under nearly physiological conditions, it forms a tubular crystal forming of rounded sheet-like two-dimensional crystals (Figure 2). At present, however, X-ray crystallography remains unable to analyze crystals other than three-dimensional crystals. In the case of thin two-dimensional crystals and tubular crystals, analysis is impossible due to the very limited availability of diffraction data. Use of nuclear magnetic resonance (NMR), a technique not requiring crystallization, is also producing significant results for the structural analysis of proteins. It should be noted, however, that because the size of the protein analyzable by NMR depends on the intensity of the magnetic field applied, its application remains limited to small molecules up to about 30,000 in molecular weight and portions (functional domains, etc.) of molecules. Because of the molecular weight of the acetylcholine receptor - approximately 290,000 - its overall structure cannot be analyzed by NMR. Hence, Miyazawa and colleagues employed a third approach known as electron crystallography, based on electron microscopy. Electron beams are more interactive with substances than X-rays. Therefore, electron beams provide robust diffraction data, enabling analysis of two-dimensional crystals and tubular crystals. However, their use is problematic in that the protein structure is destroyed by the heat from the strong interaction. To overcome this drawback, the sample must be cooled to the maximum possible extent. Miyazawa made the most use of the ultra-low-temperature electron microscope that Group Director Fujiyoshi had developed after long and painstaking work, to take electron micrographs of the acetylcholine receptor. Currently, only a few units of this type are available in the world, which are capable of taking electron micrographs while minimizing sample damage under cooling conditions at an ultra-low temperature of -269°C using liquid helium. However, it took as long as eight years to obtain about 10,000 electron micrographs needed for the structural analysis of the acetylcholine receptor. Miyazawa explains the reasons as follows: "To assign an amino acid sequence (primary structure) to a three-dimensinal structure, a resolution on the atomic scale of 4 angstroms (  ) (1 is one 10-billionth of a meter) is necessary. To achieve this level, diffraction data must be collected from as many as around one million protein molecules, and averaged. In analyzing the three-dimensional structure of an amino acid crystal using X-rays, only a single shot provides a resolution of 4 or more, sufficient to determine the atomic structure, because the crystal contains at least one hundred trillion molecules. In contrast, a tubular crystal of the acetylcholine receptor contains only 3,000 molecules; therefore, more than 300 electron micrographs must be taken. Moreover, not all photos taken serve this purpose. For example, if the tube is curved, the photos are useless for the analysis. Out of 10,000 photos, 359 proved to be useful for the analysis, and using them, we succeeded in determining the atomic structure of the acetylcholine receptor (Figure 3)."
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| Regulation with precision of one 10-billionth of a meter | |||
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What was elucidated in the functional mechanism of the acetylcholine receptor from its three-dimensional structure? The acetylcholine receptor comprises five circularly arranged subunits that penetrate the cell membrane. In the extracellular domain, there are two ligand-binding legions, where acetylcholine molecules are trapped. The channel which is the path of ions is located inside the transmembrane region (Figure 4), with five helical structures (a-helices) arranged circularly to form a pentagonal ring (Figure 5). Upon the binding of acetylcholine to the two ligand-binding regions, the regions rotate and two of the a-helices in the transmembrane region rotate synchronously. Subsequently, the pentagonal ring deforms and expands outward. Then the channel opens, followed by the influx of sodium ions. The sodium ion is 2 in diameter, whereas the ring pore is 6 even when the channel is closed. Then, why is the sodium ion unable to pass through the 6 -ring? "In the aqueous cytoplasm of the cell, sodium ions are positively-charged and are in a hydrated state bound with water molecules around. Because a hydrated sodium ion is 8 in diameter, it cannot pass through the 6 -ring. When the channel opens, however, the ring diameter increases to 9 to allow the hydrated 8 -sodium ion to pass through. Furthermore, when the channel is closed, hydrophobic amino acids such as valine and leucine are present on the ring surface and repel water. When the channel opens, however, the a-helices rotate to expose serine and other hydrophilic amino acids onto the ring surface, which in turn facilitates the passage of hydrated sodium ions." This exquisite mechanism of the receptor had not been clarified until the three-dimensional structure was determined on the atomic scale.The acetylcholine receptor was the first neurotransmitter receptor to have its structure solved at an atomic resolution. The receptor is of the type known as a "ligand-gated ion channel," comprising an integrated body of the ligand-binding regions and the ion channel. Receptors of neurotransmitters such as serotonin, GABA (g-aminobutyric acid) and glycine are of the same type, and are considered to share the same functional mechanism. The present achievements of Miyazawa and his colleagues will no doubt make a significant contribution to brain science and neuroscience.
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| Elucidating the etiology | |||
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"The first telephone call I received after publication of the present work was from a doctor at a medical school. He said, 'Your achievement will enable the elucidation of the etiology of epilepsy , which my patients are suffering from!'" Epilepsy is a disease characterized by specific symptoms such as seizures, convulsions, and loss of consciousness due to the abnormal activities of neurons in the brain. The amino acid substitution of serine by leucine in a portion of the neurotransmitter receptor, caused by a genetic mutation, has been found in patients suffering from one form of familial epilepsy. However, it remained unknown how the substitution is associated with receptor functions. "As we found, normally, when the acetylcholine receptor channel opens, the a-helices rotate to expose a hydrophilic amino acid, serine, onto the surface of the receptor, and this makes the passage of hydrated ions easier, whereas actually the serine is replaced by leucine in familial epilepsy. Hence, it can be considered that even when the channel opens, it does not allow the ions to pass through." The achievement of Miyazawa and colleagues has also opened a way to elucidating the etiology of myasthenia, a disease characterized by progressive loss of muscle strength. "In familial myasthenia, an amino acid variation was found at the junction between the receptor's ligand-binding region and the transmembrane portion. It is suspected that even when acetylcholine has bound and the ligand-binding region has rotated, concerted regulation of the transmembrane region fails and channel gating cannot be controlled." Determining the three-dimensional structures of proteins leads directly to the elucidation of the etiology of disease, and opens a way to the radical treatment of intractable diseases. "Even if the protein structure is clarified from our work, it does not mean that it makes intractable diseases treatable immediately. However, the answer to the question of why 'things have come to this' could be provided for patients and physicians struggling with diseases. I hope that human beings will see the accumulation of persuasive scientific evidence as of the utmost importance." |
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| New analytical methods based on electron microscopy | |||
 Miyazawa is now working to develop sophisticated analytical methods based on electron microscopy. The yellow portion in Figure 6 shows the overall structure of the protein known as the IP3 receptor as determined by single-particle analysis. "The three-dimensional structure was obtained from the electron micrograph of each purified protein taken from various directions. In this method, the subject protein need not be crystallized, but the position of amino acid residues in the structure cannot be determined due to its poor resolution of 10 to 20 . If the whole structure is known, it will be possible to fit the partial structures, determined at atomic resolution using other analytical methods, to the unanalyzed parts. However, this analytical method also has not been completed yet. Our success must be attributed to the prophetic vision of Dr. Katsuhiko Mikoshiba, group director of the Brain Science Institute at RIKEN and Professor at the University of Tokyo, who discovered the IP3 receptor and lead this study, to the efforts of Dr. Chikara Sato of the National Institute of Advanced Industrial Science and Technology, who worked with us in part of analysis, and to the superb vision of the leader of this group, Prof. Fujiyoshi. This is really an 'all-out war' as described by Prof. Fujiyoshi, rather than joint research."The research team is also engaged in developing a new technique called "electron tomography." "In the cell, proteins exhibit their functions while interacting with many other proteins. It is necessary to know the arrangement of proteins and their interaction in tissue for an understanding of protein functions. It is possible to know the 3D arrangement of proteins at a resolution of 30 to 50 by photographs of tissue sections taken from various directions." However, such images cannot tell us the difference between the sorts of proteins. Miyazawa and colleagues are working to develop a technology for accurately incorporating a label observable under an electron microscopy into an randomly-chosen protein."There are three approaches to protein analysis by electron microscopy: electron crystallography, which explores structures on the atomic scale; single-particle analysis, which provides data on the whole structure of a single molecule; and electron tomography, which visualizes a group of proteins as embedded in tissue. Analysis using these three methods in combination should bring to light how proteins work in the body to support the mechanism of life." |
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| Aiming at structural medicine | |||
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Miyazawa emphasizes, "To date easily-analyzable proteins using either X-rays or electron beams have been subjected to structural determination, but this practice is behind the times now." Now, most of the proteins whose three-dimensional structures have been determined are bacterial proteins, which produce many molecules and have crystals that are easy to obtain. "Traditionally, the common approach has been to search and analyze the bacterial proteins that are similar to those acting in the human body. However, they are not identical to human proteins. Currently, thanks to the advances in molecular biology, it is possible to express a human gene to produce many protein molecules, and to obtain their crystals." Miyazawa is about to launch a new project aiming for structural medicine, which will be established by advancing electron microscopy techniques for structural analysis to help improve medical treatment. "We have received messages from a number of physicians struggling with intractable disease, saying that they were much encouraged by our discovery of the structure of the acetylcholine receptor. Even basic research like ours can help promote human welfare. The time has come for us to be active in investigating those proteins that must be analyzed, i.e., proteins associated with human diseases, rather than those that are just easily analyzable."
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