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3D Visualization and Quantification of Bioplastic PHA in a Living Bacterial Cell
3D holographic microscopy leads to in-depth analysis of bacterial cells accumulating the bacterial bioplastic, polyhydroxyalkanoate (PHA) A research team at KAIST has observed how bioplastic granule is being accumulated in living bacteria cells through 3D holographic microscopy. Their 3D imaging and quantitative analysis of the bioplastic ‘polyhydroxyalkanoate’ (PHA) via optical diffraction tomography provides insights into biosynthesizing sustainable substitutes for petroleum-based plastics. The bio-degradable polyester polyhydroxyalkanoate (PHA) is being touted as an eco-friendly bioplastic to replace existing synthetic plastics. While carrying similar properties to general-purpose plastics such as polyethylene and polypropylene, PHA can be used in various industrial applications such as container packaging and disposable products. PHA is synthesized by numerous bacteria as an energy and carbon storage material under unbalanced growth conditions in the presence of excess carbon sources. PHA exists in the form of insoluble granules in the cytoplasm. Previous studies on investigating in vivo PHA granules have been performed by using fluorescence microscopy, transmission electron microscopy (TEM), and electron cryotomography. These techniques have generally relied on the statistical analysis of multiple 2D snapshots of fixed cells or the short-time monitoring of the cells. For the TEM analysis, cells need to be fixed and sectioned, and thus the investigation of living cells was not possible. Fluorescence-based techniques require fluorescence labeling or dye staining. Thus, indirect imaging with the use of reporter proteins cannot show the native state of PHAs or cells, and invasive exogenous dyes can affect the physiology and viability of the cells. Therefore, it was difficult to fully understand the formation of PHA granules in cells due to the technical limitations, and thus several mechanism models based on the observations have been only proposed. The team of metabolic engineering researchers led by Distinguished Professor Sang Yup Lee and Physics Professor YongKeun Park, who established the startup Tomocube with his 3D holographic microscopy, reported the results of 3D quantitative label-free analysis of PHA granules in individual live bacterial cells by measuring the refractive index distributions using optical diffraction tomography. The formation and growth of PHA granules in the cells of Cupriavidus necator, the most-studied native PHA (specifically, poly(3-hydroxybutyrate), also known as PHB) producer, and recombinant Escherichia coli harboring C. necator PHB biosynthesis pathway were comparatively examined. From the reconstructed 3D refractive index distribution of the cells, the team succeeded in the 3D visualization and quantitative analysis of cells and intracellular PHA granules at a single-cell level. In particular, the team newly presented the concept of “in vivo PHA granule density.” Through the statistical analysis of hundreds of single cells accumulating PHA granules, the distinctive differences of density and localization of PHA granules in the two micro-organisms were found. Furthermore, the team identified the key protein that plays a major role in making the difference that enabled the characteristics of PHA granules in the recombinant E. coli to become similar to those of C. necator. The research team also presented 3D time-lapse movies showing the actual processes of PHA granule formation combined with cell growth and division. Movies showing the living cells synthesizing and accumulating PHA granules in their native state had never been reported before. Professor Lee said, “This study provides insights into the morphological and physical characteristics of in vivo PHA as well as the unique mechanisms of PHA granule formation that undergo the phase transition from soluble monomers into the insoluble polymer, followed by granule formation. Through this study, a deeper understanding of PHA granule formation within the bacterial cells is now possible, which has great significance in that a convergence study of biology and physics was achieved. This study will help develop various bioplastics production processes in the future.” This work was supported by the Technology Development Program to Solve Climate Changes on Systems Metabolic Engineering for Biorefineries (Grants NRF-2012M1A2A2026556 and NRF-2012M1A2A2026557) and the Bio & Medical Technology Development Program (Grant No. 2021M3A9I4022740) from the Ministry of Science and ICT (MSIT) through the National Research Foundation (NRF) of Korea to S.Y.L. This work was also supported by the KAIST Cross-Generation Collaborative Laboratory project. -PublicationSo Young Choi, Jeonghun Oh, JaeHwang Jung, YongKeun Park, and Sang Yup Lee. Three-dimensional label-free visualization and quantification of polyhydroxyalkanoates in individualbacterial cell in its native state. PNAS(https://doi.org./10.1073/pnas.2103956118) -ProfileDistinguished Professor Sang Yup LeeMetabolic Engineering and Synthetic Biologyhttp://mbel.kaist.ac.kr/ Department of Chemical and Biomolecular Engineering KAIST Endowed Chair Professor YongKeun ParkBiomedical Optics Laboratoryhttps://bmokaist.wordpress.com/ Department of PhysicsKAIST
Meet the KAISTian of 2017, Professor YongKeun Park
Professor YongKeun Park from the Department of Physics is one of the star professors in KAIST. Rising to the academic stardom, Professor Park’s daily schedule is filled with series of business meetings in addition to lab meetings and lectures. The year 2017 must have been special for him. During the year, he published numerous papers in international journals, such as Nature Photonics, Nature Communications and Science Advances. These high performances drew international attention from renowned media, including Newsweek and Forbes. Moreover, recognizing his research performance, he was elected as a fellow member of the Optical Society (OSA) in his mid-30s. Noting that the members’ age ranges from late 50s to early 60s, Professor Park’s case considered to be quite exceptional. Adding to his academic achievement, he has launched two startups powered of his own technologies. One is called Tomocube, a company specialized in 3-D imaging microscope using holotomography technology. His company is currently exporting the products to multiple countries, including the United States and Japan. The other one is The.Wave.Talk which has technologies for examining pre-existing bacteria anywhere and anytime. His research career and entrepreneurship are well deserved recipient of many honors. At the 2018 kick-off ceremony, Professor Park was awarded the KAISTian of 2017 in recognition of his developing holographic measure and control technology as well as founding a new field for technology application. KAISTian of the Year, first presented in 2001, is an award to recognize the achievements and exemplary contribution of KAIST member who has put significant effort nationally and internationally, enhancing the value of KAIST. While receiving the award, he thanked his colleagues and his students who have achieved this far together. He said, “I would like to thank KAIST for providing environment for young professors like me so that we can engage themselves in research. Also, I would like to mention that I am an idea seeder and my students do the most of the research. So, I appreciate my students for their hard works, and it is very pleasure to have them. Lastly, I thank the professors for teaching these outstanding students. I feel great responsibility over this title. I will dedicate myself to make further progress in commercializing technology in KAIST.” Expecting his successful startup cases as a model and great inspiration to students as well as professors, KAIST interviewed Professor Park. Q What made you decide to found your startups? A I believed that my research areas could be further used. As a professor, I believe that it is a university’s role to create added value through commercializing technology and creating startups. Q You have co-founded two startups. What is your role in each company? A So, basically I have two full-time jobs, professor in KAIST and CTO in Tomocube. After transferring the technology, I hold the position of advisor in The.Wave.Talk. (Holographic images captured by the product Professor Park developed) Q Do your students also participate in your companies or can they? A No, the school and companies are separate spaces; in other words, they are not participating in my companies. They have trained my employees when transferring the technologies, but they are not directly working for the companies. However, they can participate if they want to. If there’s a need to develop a certain technology, an industry-academia contract can be made. According to the agreement, students can work for the companies. Q Were there any hardships when preparing the startups? A At the initial stage, I did not have a financial problem, thanks to support from Startup KAIST. Yet, inviting capital is the beginning, and I think every step I made to operate, generate revenue, and so on is not easy. Q Do you believe KAIST is startup-friendly? A Yes, there’s no school like KAIST in Korea and any other country. Besides various programs to support startup activities, Startup KAIST has many professors equipped with a great deal of experience. Therefore, I believe that KAIST provides an excellent environment for both students and professors to create startups. Q Do you have any suggestion to KAIST institutionally? A Well, I would like to make a comment to students and professors in KAIST. I strongly recommend them to challenge themselves by launching startups if they have good ideas. Many students wish to begin their jobs in government-funded research institutes or major corporates, but I believe that engaging in a startup company will also give them valuable and very productive experience. Unlike before, startup institutions are well established, so attracting good capital is not so hard. There are various activities offered by Startup KAIST, so it’s worthwhile giving it a try. Q What is your goal for 2018 as a professor and entrepreneur? A I don’t have a grand plan, but I will work harder to produce good students with new topics in KAIST while adding power to my companies to grow bigger. By Se Yi Kim from the PR Office
Professor YongKeun Park Elected as a Fellow of the Optical Society
Professor YongKeun Park, from the Department of Physics at KAIST, was elected as a fellow member of the Optical Society (OSA) in Washington, D.C. on September 12. Fellow membership is given to members who have made a significant contribution to the advancement of optics and photonics. Professor Park was recognized for his research on digital holography and wavefront control technology. Professor Park has been producing outstanding research outcomes in the field of holographic technology and light scattering control since joining KAIST in 2010. In particular, he developed and commercialized technology for a holographic telescope. He applied it to various medical and biological research projects, leading the field worldwide. In the past, cells needed to be dyed with fluorescent materials to capture a 3-D image. However, Professor Park’s holotomography (HT) technology can capture 3-D images of living cells and tissues in real time without color dyeing. This technology allows diversified research in the biological and medical field. Professor Park established a company, Tomocube, Inc. in 2015 to commercialize the technology. In 2016, he received funding from SoftBank Ventures and Hanmi Pharmaceutical. Currently, major institutes, including MIT, the University of Pittsburgh, the German Cancer Research Center, and Seoul National University Hospital are using his equipment. Recently, Professor Park and his team developed technology based on light scattering measurements. With this technology, they established a company called The Wave Talk and received funding from various organizations, such as NAVER. Its first product is about to be released. Professor Park said, “I am glad to become a fellow member based on the research outcomes I produced since I was appointed as a professor at KAIST. I would like to thank the excellent researchers as well as the school for its support. I will devote myself to continuously producing novel outcomes in both basic and applied fields.” Professor Park has published nearly 100 papers in renowned journals including Nature Photonics, Nature Communications, Science Advances, and Physical Review Letters.
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