A useful substance discovered in a nuisance creature
16 November 2007 (Volume 2 Issue 11)
The Echizen, or Nomura’s, jellyfish () has a giant bell exceeding one meter across, weighs more than 100 kg, and is the largest of its kind in the world. Recently it has overpopulated the Sea of Japan, causing major damage to fisheries. Kiminori Ushida, Research Unit Leader of the Eco-Soft Materials Research Unit at the RIKEN Discovery Research Institute, and colleagues found that this nuisance creature contains large amounts of a useful substance, attracting a lot of attention. The substance is a type of glycoprotein generically known as mucin, which has antibacterial and moisture-retaining properties, among others. Ushida and colleagues are working to modify the sugar chain of this new mucin to enhance its function so that it can be used in a new drug.
The discovery of the valuable biomaterial, mucin
Why did Ushida take note of jellyfish?
Ushida’s Eco-Soft Material Research Unit is investigating soft matters, including polymers and biomolecules, with the aim of contributing to society, particularly with respect to environmental issues. “I have been engaged in research into hyaluronic acid (hyaluronan), a saccharide polymer, for some 10 years,” he says. Hyaluronic acid has recently become popular as an ingredient for cosmetics and other products. Ushida is also researching the disposal of waste plastics. “I have been aware of the fact that people were troubled over how to legally dispose of jellyfish clogging water intakes at power plants,” he explains. When he was informed of the unexpectedly low protein content of the jellyfish, it occurred to him that they might be rich in saccharides. “This motivated me to look more closely at jellyfish, although I first focused on the extraction of hyaluronic acid.”
Ushida started his academic career as a researcher in basic physical chemistry and this remains an area of interest to him “I did not have a good knowledge of the biochemical technology needed to extract the desired components,” says Ushida. “So I bought an introductory text [book] and made experiments, consulting it frequently.” He says that the method of extraction he used was quite simple—at the level of university bachelor students. In this way, he successfully obtained many precipitates. “I asked Dr. Naoshi Dohmae, Team Leader of the Biomolecular Characterization Team at the RIKEN Advanced Development and Supporting Center, to analyze them.” He explains that this is common at RIKEN; researchers find it easy to ask scientists in other laboratories for their cooperation. Dohmae willingly consented to undertake the analytical work, although he said, “Further purification would be appreciated before his task.”
Hence in December 2004, the amino acid sequence of the extract was determined and the substance was identified as mucin, a member of the glycoprotein family (Fig 1).
Figure 1: Qniumucin under purification.
Because qniumucin is freely soluble in water and can be easily prepared as a membrane, it is expected to serve as a raw material for new biodegradable materials (medical films, etc.).
Ushida’s research was accelerated by his associates. He was working in the laboratory of the late Hiroyuki Hatano, then professor at Kyoto University. Hatano was famous worldwide as the inventor of the prototype of an amino acid analyzer. “In those days, I was under the direct guidance of associate professor Tadamasa Shida. Kazuyuki Akasaka—now a professor at Kinki University and a pioneer of protein structural analysis by nuclear magnetic resonance (NMR)—served as an assistant staff in the laboratory,” he recalls. “Hence, my acquaintances include many experts in amino acid analysis or NMR structural analysis. Relying on their advice, I was able to continue to analyze the new mucin and nearly completed the structural analysis.”
“How can we handle the monsters?”
To start the investigation, a great many jellyfish were required. “I was referred to the Mitsu Fisheries Cooperative Association by my old acquaintance at Kyotango City in Kyoto Prefecture, where I used to go swimming every year when I was a member of Hatano’s laboratory,” says Ushida. “It was in May 2005. One chief skipper there asked me, ‘Can you find any good idea for handling these monsters?’”
The “monsters” refer to Nomura’s jellyfish (Fig. 2). Up to that time the water (or moon) jellyfish (Aurelia aurita) had been the major study material for Ushida and colleagues. Its bell measures at most 10 cm to 20 cm across. Nomura’s jellyfish, with a bell exceeding one meter in diameter and weighing more than 100 kg, are much larger (Fig. 3). Do these monsters also contain mucin?
Figure 2: Nomura’s jellyfish (Nemopilema nomurai) on the float.
Qniumucin has been found in nearly all species of jellyfish occurring in the seas around Japan. Qniumucin accounts for 0.02 to 0.1% by weight of the whole body of the jellyfish.
In August 2005, Ushida filed a patent application for the new mucin, and soon afterwards, there was an outbreak of Nomura’s jellyfish. “I asked Shinwa Chemical Industries Ltd in Kyoto City, with whom I already had research links, to extract some mucin, and finally the extract was analyzed at RIKEN.” Surprisingly, Nomura’s jellyfish was found to contain a mucin with exactly the same amino acid sequence as the moon jellyfish. “From there, I examined a variety of jellyfish species living in the seas around Japan and found the same mucin with the same amino acid sequence in almost all of them.”
Around that time, Ushida began considering commercializing mucin extraction. He describes how, when removed from the sea, jellyfish form no more than a virtually useless waste to be disposed of. If mucin can be extracted during the disposal with a sufficient yield to allow industrial production, manufacturing, and marketing, the benefits could be returned to the local people suffering from these nuisances. “I named this new mucin ‘qniumucin’ after the kuniumi shinwa (birth-myth of the nation of Japan) as told in Kojiki (Records of Ancient Matters) with the expectation of contributing to regional economical development.”
Figure 3: Numerous Nomura’s jellyfish entangled in a stationary net (off the coast of Amino Town in Kyotango City).
Recently, the surge of Nomura’s jellyfish in the Sea of Japan amounts to several thousands to several tens of thousands of tons even from one beach, causing major damage to local fisheries. Once removed from the sea, the jellyfish must be disposed of in the most effective way as industrial waste. It seems most effective to remove the water, which accounts for about 95% of its body weight, then dispose of the residual solid matter separately. Qniumucin can easily be extracted concurrently with the disposal treatment.
Although some animal mucin products, including preparations from the gastric juices of cattle, are commercially available, their applications are limited to use as a food additive and cosmetic ingredient. Amid this situation, particularly with respect to the recently arising concerns regarding bovine spongiform encephalopathy (BSE), there is a strong demand for the development of highly safe animal mucins.
Ushida is now engaged in developing a manufacturing process and business model for qniumucin in preparation for launching a venture enterprise. The key to business success resides in expanding the uses of qniumucin to bring increased value.
Medical applications
There are numerous types of mucin, including those found in animal digestive juices such as gastric juices and saliva, as well as in the mucous membranes of the nose and eye. About 12 types have been identified in the human body. In addition to animal sources, mucins also form the major ingredients of the gooey substances of okra and yam plants.
A mucin molecule consists of a protein moiety of amino acids in a sequence and a sugar chain moiety of sugar molecules in a chain. For example, the mucin contained in the mucus adsorbs bacteria and viruses, suppressing their activity and also acting as a surfactant that incorporates them in the mucus and washes them away. In this process, the sugar chain of the mucin is said to recognize and adsorb the proteins on the surfaces of the bacteria and viruses.
The amino acid sequences for the types of mucin found in the human body have been determined, but how the sugar molecules join together to form the sugar chains remains unknown. This is because of the technical difficulties involved in examining sugar chain structures, which are more complex than protein structures. The structures of the protein and sugar chain have not yet been completely clarified for any type of mucin.
“For qniumucin, however, we have succeeded in elucidating more than 90% of the structure of its sugar chain, as well as the amino acid sequence of its protein,” says Ushida (Fig. 4). “This is thanks to the structural simplicity of qniumucin.”
Figure 4: Tandem repeats and molecular structures of qniumucin.
Mucin is a generic name for the family of polymer compounds comprising simple tandem repeats of amino acids and sugar chains bound thereto via O-glycoside bonds. Qniumucin and MUC5AC (a human mucin) have tandem repeats of eight amino acids, of which four are common to both. Ushida and colleagues have nearly clarified the primary structure of qniumucin (right).
The qniumucin protein is not complicated; it consists of simple repeats of eight amino acids. “In addition, the mucin found in the human stomach and esophagus, MUC5AC, also consists of repeats of eight amino acids, of which four are shared by qniumucin; they have very high structural similarity,” says Ushida (Fig. 4). Hence, even if qniumucin is used in the human body, it is unlikely to cause rejection reactions or allergies due to immune-system responses that eliminate foreign substances.
“The most likely applications of qniumucin are artificial gastric juices, artificial saliva, and eye drops,” says Ushida. “Many elderly people suffer from a lack of salivation. I am investigating the potential use of qniumucin for artificial saliva, in consultation with a researcher based in a university department of dentistry.” Essentially, ordinary mucin has no special potent physiological action. However,a preparation of a new mucin, that is structurally similar to a human mucin, and that is artificially produced from qniumucin would find new applications as artificial saliva and the like.
Ushida is also conducting research to apply qniumucin in the treatment of another disease affecting many patients. He says, “Unfortunately, the facts have not yet been laid before the public.” In the near future, the promising results of his applied research into qniumucin will be announced.
Designing a new mucin
Qniumucin has a simple sugar chain structure, and can be obtained from a variety of jellyfish, as a unique substance with the constant amino acid sequence and the constant sugar chain structure. Ushida says this is the most appealing feature of qniumucin. Qniumucin can now be handled as a subject of chemical research because its steric molecular structure has been almost fully determined. A new discipline—mucin chemistry—is ready to be launched. “We aim to design new mucins using qniumucin as the starting material.”
Although artificial synthesis of mucin is being studied, the goal seems to be far away. Its protein moiety, biosynthesized on the basis of genetic information in the body, can be created by means of genetic engineering and other techniques, but its sugar chain cannot be synthesized successfully. “However, some glycotransferases have been discovered that replace sugar molecules with others. I would like to use them to take on the task of creating mucins that have new functions by altering the sugar chain of qniumucin,” says Ushida. He explains that, for example, the mucin contained in the mucus is not highly potent for the adsorption of bacteria and viruses, but if its sugar chain is modified to enhance its adsorption power, the mucin may find a pharmaceutical application.
The discovery of qniumucin was announced in the Journal of Natural Products, published jointly by the American Chemical Society (ACS) and the American Society of Pharmacognosy, on June 23, 2007. The ACS selected the article as a noteworthy paper for press release, and the article created a major sensation in the United States and Europe, where people are highly conscious of environmental issues. The story of the discovery also featured in daily newspapers, including the New York Times in the USA and the Daily Telegraph in the UK. This global interest is due to an explosion in the jellyfish population, which is occurring in seas all over the world.
“The American Chemical Society’s press release suggests the potential for designer mucins, and provides a good summary of what I think is scientifically most important,” says Ushida. US researchers will soon begin studying how to design new mucins. “We must not get left behind,” he adds. “Seeking cooperation from various researchers at RIKEN once more, we have already commenced a study to modify the sugar chain of qniumucin.”
Comprehensive and profound understanding—the ultimate goal of researchers in basic sciences
“A big advantage of RIKEN resides in the fact that top researchers in a broad range of fields are available at one’s elbow, and so one can establish multidisciplinary friendships,” says Ushida. Ushida believes that researchers in basic sciences must not confine themselves to their own expertise. “Ask an expert in the relevant field what you cannot understand, and proceed with determination—I think this is the essential approach, although such tenacious researchers, like me, may form a small minority.”
“The ultimate goal of the basic sciences is to know the essences of nature. To this end, a broad range of events must be understood both comprehensively and profoundly. In research into qniumucin, for example, we cannot be successful unless we fully understand NMR data and sugar chain synthesis, which were once out of our expertise. One important role of researchers in the basic sciences is to derive leading principles that underlie the technologies being put into practical application. To achieve this, it is essential to understand nature, both comprehensively and profoundly, while having personal exchanges with colleagues in a broad range of fields.”
(Interview/writer in Japanese: Akira Tateyama)
About the Researcher
Kiminori Ushida
Kiminori Ushida was born in Osaka, Japan, in 1958. He graduated from the Faculty of Science, Kyoto University, in 1982, and obtained his PhD in 1987 from the same university in radiation chemistry. He spent three years as an assistant professor at the Kyoto Institute of Technology, Japan, where he began studying ultrafast laser spectroscopy. He joined RIKEN as a research scientist in the Chemical Dynamics Laboratory, where he was involved with three research groups for Solar Science and Nano Science working on several projects dealing with nonlinear spectroscopy. He was promoted to senior scientist in 1996 and to research unit leader in 2004. Since then, he has been director of his own research group for soft material study of which applications to environmental issues are in his scope. His research area covers a very wide range of subjects, for example: basic chemical physics on extracellular matrices using fluorescence correlation spectroscopy, a developmental study on the treatment of plastic wastes by microwave, and this study about the novel mucin extracted from jellyfishes.
