2nd North America – Japan Enzyme Technology Symposium Speakers

Satoru Ishihara, Ph.D.

• General Manager
• Frontier Research Department
• Innovation Division
• Amano Enzyme Inc.
• [email protected]

Biography

Satoru Ishihara received his Ph.D. in Biology from the University of Tokyo in 2005. He then worked as a postdoctoral fellow at two universities in Japan. In 2010, he joined Amano Enzyme Inc., focusing on the development of industrial enzymes. From 2015 to 2017, he served as a scientist at Amano Enzyme USA Co., Ltd. in Elgin, IL. Since returning to Japan in 2017, he has held management roles, including Assistant Team Leader from 2020 to 2022, Team Leader from 2022 to 2024, and General Manager of Frontier Research Department since 2024.

Research Interests

• Developments of industrial enzymes.
• Enzyme engineering
• Advanced technologies for enzyme design

Abstract

Performance expansion of industrial specialty enzymes

Satoru Ishihara

The use of enzymes as environmentally friendly and sustainable alternatives is expanding across diverse industrial processes. According to the expanded application, various enzymes have been sourced from both nature and in silico database searches. Their properties are being enhanced by protein engineering. Amano Enzyme Inc. has succeeded in developing and improving industrial enzymes by combination of conventional and computational approaches. This presentation will describe enzymes, which were originally found from nature, have been improved using several protein engineering methods, including rational design, directed evolution, and machine learning-assisted approaches.

Protein-glutaminase (PG) from Chryseobacterium proteolyticum has the catalytic activity to deamidate the amide groups of glutamine residues in proteins, converting them into glutamic acid and releasing ammonia. In the food industry, PG is used to improve the physical properties and texture of plant-based foods. We have successfully isolated PG-producing bacteria from soil and commercialized the product. [1, 2] Through protein engineering, we have developed several PG mutants with significantly improved properties, including thermal stability and activity at higher temperature. On-going, we are investigating how to find combinations with various mutations efficiently using machine learning models.

Lipase (BCL) from Burkholderia cepacia is used in the production of fragrances and bioactive compounds. BCL is commercialized as Lipase PS “Amano” from Amano Enzyme Inc. and is one of the most useful biocatalysts. BCL has been reported to catalyze the transformation of racemates into single enantiomers. In collaboration with National Institute of Advanced Industrial Science and Technology, we used molecular dynamics simulations to enhance the enantioselectivity. The engineered BCLs have enhanced their enantioselectivity for target products, improving the production purity, and have been promising enzymes for large-scale bioprocesses. [3]

References:

[1] S. Yamaguchi et al., Appl. Environ. Microbial., 2000, 66, 3337

[2] S. Yamaguchi et al., Eur. J. Biochem., 2001, 268, 1410 

{3} J. Ikebe, et al., Journal of Agricultural and Food Chemistry 2025, 73, 8, 4829.

Kate Adamala, Ph.D. 

• McKnight Presidential Fellow Associate Professor,
• University of Minnesota
• [email protected]

Biography

Kate Adamala received her PhD in biophysics from the University Roma Tre. Her research aims at understanding chemical principles of biology, using artificial cells to create new tools for bioengineering, drug development, and foundational research. Her research interests span questions from the origin and earliest evolution of life, using synthetic biology to colonize space, to the future of biotechnology and medicine. 

Kate is a co-founder of the synthetic cell therapeutics startup Synlife, a leader of the BioBOLD Initiative, and co-founder and coordinator of the international synthetic cell engineering consortium Build-a-Cell. Lab info protobiology.org.

Research Interests

• Synthetic cells.
• Bioengineering
• Introduction of advanced technologies into enzyme design
• Application of digital biotechnologies to fermentation process 

Abstract

Bioengineering with synthetic cells

Kate Adamala

All of the current biological research is done on a single sample: we know only one type of modern terrestrial life. By engineering synthetic living systems, we seek to expand that sample size, exploring properties of lineage agnostic organisms.

Synthetic cells are liposomal bioreactors that have some, but not all properties of live cells. Creating artificial living systems allows us to diversify the chassis of biological studies, and provides novel opportunities for bioengineering. We can begin to answer questions about healthy and diseased natural cells, and ask new questions about the limits of biology. Engineering synthetic cells with fundamentally different physical and chemical properties, we can compare behaviors and begin drawing broad conclusions about basic rules of biological life.

Synthetic cells are fully defined and engineerable, enabling studies of natural processes with level of detail unavailable in complex and messy live cell. In synthetic cell systems, there is less noise from underlying endogenous processes of central metabolism, and no evolutionary limitations of survivability. Synthetic cells provide new chassis for biological studies, for broadening understanding of our own type of biology, and for investigating alternatives to the single known life form.

Hal Alper, Ph.D. 

• Cockrell Family Regents Chair in Engineering
• The University of Texas at Austin
• [email protected]

Biography

Dr. Alper earned his Ph.D. in Chemical Engineering from MIT (2006). His research focuses on enabling sustainable biotechnology through applying the approaches of metabolic engineering, synthetic biology, systems biology, and protein engineering. Dr. Alper has published over 175 articles and 8 book chapters. He is a fellow to the American Institute for Medical and Biological Engineering (2018), the National Academy of Inventors (2019), and the American Association for the Advancement of Science (2024).

Research Interests

• Enzymatic and cellular approaches to biocatalysis
• Protein engineering and directed evolution approaches
• Metabolic Engineering and Synthetic Biology

Abstract

Enzymatic depolymerization of plastics

Hal Alper

Plastic waste is an environmental challenge, but also presents a biotechnological opportunity as a unique carbon substrate. With modern biotechnological tools, it is possible to enable both recycling and upcycling. To realize a plastics bioeconomy, significant intrinsic barriers must be overcome using a combination of enzyme, strain, and process engineering. This presentation will highlight the use of bioprospecting, bioinformatics, structure-based machine learning, and enzyme engineering to identify novel enzyme catalysts for the depolymerization of various plastics including PET, PE, and PHB. Considerations for practical use and deployment will be discussed. Finally, recent advances and prospects for other plastics will also be described as a window into the future for establishing a plastics bioeconomy.

Saori Kosono, Ph.D. 

• Associate Professor
• Graduate School of Agriculture and Life Sciences;
• Collaboration Research Institute for Innovative Microbiology
• The University of Tokyo
• [email protected]

Biography

Saori Kosono received Ph.D. degrees from Osaka University (1996). Her doctoral thesis focused on plasmid conjugation in Streptomyces species. She joined RIKEN, Japan, as a postdoctoral researcher (1996–1997), and subsequently held positions as a research scientist (1997–2005) and senior research scientist (2005–2012). She has been conducting research on bacterial adaptation, as well as post-translational modifications and metabolic regulation in bacteria. In 2012, she was appointed as a project associate professor at the University of Tokyo, and in 2019, she became an associate professor. She received the Award for Women Scientists from the Japan Society for Bioscience, Biotechnology, and Agrochemistry in 2018.

Research Interests

• Post-translational modifications
• Metabolic regulation
• Protein condensation

Abstract

Catalysis-dependent condensation of citrate synthase involved in glutamate overproduction in Corynebacterium glutamicum

Saori Kosono

Recent studies suggest that protein condensation plays a role in regulating cellular metabolism. Corynebacterium glutamicum, a gram-positive bacterium with high GC content, is generally recognized as safe (GRAS) and is widely used for industrial production of amino acids, particularly L-glutamate (an umami component). In C. glutamicum, overproduction of extracellular glutamate induces a global increase in metabolic flux toward glutamate synthesis without a corresponding increase in metabolic enzyme levels, implying the involvement of post-translational regulatory mechanisms. We aimed to elucidate the post-translational mechanisms underlying global metabolic changes associated with glutamate overproduction and discovered that citrate synthase (CgCS) formed droplet-like condensates in C. glutamicum cells. CgCS condensation was observed in growing cells containing catalytically active CgCS. In vivo and in vitro studies revealed that oxaloacetate, a substrate of CS, regulated condensate formation. We propose that the substrate-assisted conformational change is vital for the condensation of CgCS, that is, CgCS condensation is catalysis-dependent. Notably, the impairment of CgCS condensation was correlated with defective extracellular glutamate production. In glutamate production-deficient strains, intracellular amino acid profiles fluctuated, although the metabolism to supply 2-oxoglutarate was active. Our results suggest that CgCS condensation plays a distinct role in glutamate overproduction-related metabolism.

References

[1] M. Nagaoka, J. Yamamoto, M. Nishiyama, S. Kosono, 27 May 2025, Preprint (Ver 1) available at Research Square [https://doi.org/10.21203/rs.3.rs-6691977/v1]

Huimin Zhao, Ph.D. 

• Steven L. Miller Chair
• University of Illinois Urbana-Champaign (UIUC)
• Director
• NSF AI Institute for Molecular Synthesis
• NSF iBioFoundry
• NSF Global Center for Reliable and Scalable Biofoundries
• [email protected]

Biography

Dr. Zhao received his Ph.D. degree in Chemistry from the California Institute of Technology in 1998. Prior to joining UIUC in 2000, he was a project leader at the Industrial Biotechnology Laboratory of the Dow Chemical Company. He was promoted to full professor in 2008. Dr. Zhao has authored and co-authored over 460 research articles and over 30 issued and pending patent applications. In addition, he has given over 530 plenary, keynote, or invited lectures.

Research Interests

• Synthetic Biology
• Artificial Intelligence and Machine Learning
• Laboratory automation
• Biocatalysis and Enzyme Engineering
• Metabolic Engineering
• Natural Product Biosynthesis
• Gene Therapy 

Abstract

Expanding the Boundary of Biocatalysis: From Directed Evolution to AI-enabled Autonomous Experimentation

Huimin Zhao

Biocatalysis has been increasingly used for practical synthesis of chemicals, fuels, and materials thanks to recent advances in enzyme engineering, synthetic biology, artificial intelligence (AI)/machine learning (ML), and laboratory automation. In this talk, I will discuss our recent effort in designing novel synthetic routes and repurposed enzymes for synthesis of fine chemicals by exploring the synergy between enzymatic catalysis and non-enzymatic catalysis. Particularly, I will highlight our new strategies of combining photocatalysis with biocatalysis for abiological transformations. In addition, I will highlight the development of AI/ML and laboratory automation tools for enzyme discovery and engineering. Finally, I will introduce our new AI/ML tools for synthesis planning that integrates chemical catalysis and biocatalysis. All these strategies and tools should greatly accelerate the development of biocatalysts for applications related to human health, energy, and sustainability.  

References

[1] X. Huang, B. Wang, Y. Wang, G. Jiang, J. Feng, and H. Zhao. “Photoenzymatic Enantioselective Intermolecular Radical Hydroalkylation.” Nature, 584, 69–74 (2020). 

[2] T. Yu, A. G. Boob, M. J. Volk, X. Liu, H. Cui, and H. Zhao. “Machine learning-enabled Retrobiosynthesis of Molecules.” Nature Catalysis, 6, 137-151 (2023).

[3] T. Yu, H. Cui, J. Li, Y. Luo, G. Jiang, and H. Zhao. “Enzyme Function Prediction using Contrastive Learning.” Science, 379, 1358–1363 (2023).

[4] M. Li, Y. Yuan, W. Harrison, Z. Zhang, and H. Zhao. “Asymmetric Photoenzymatic Incorporation of Fluorinated Motifs into Olefins.” Science, 385, 416–421 (2024). 

Michelle Chang, Ph.D. 

• Professor
• Department of Chemistry
• Princeton University
• [email protected] 

Biography

Michelle received her Ph.D. degree from the Department of Chemistry at the Massachusetts Institute of Technology (2004). and carried out postdoctoral research at the University of California, Berkeley (2004-2007). She started her independent career at UC Berkeley in 2007 and recently moved to Princeton University in 2024 where is appointed as the A. Barton Hepburn Professor of Chemistry. Her group works at the interface of enzymology and biocatalysis and synthetic biology.

Research Interests

• Enzyme discovery and engineering
• Natural products biosynthesis
• Enzyme mechanism

Abstract

Discovery and engineering of new enzymes for biocatalysis

Michelle Chang

Living systems have evolved the capacity to carry out many chemical transformations of interest to synthetic chemistry if they could be redesigned for targeted purposes. Our group is interested in the discovery, characterization, and engineering of new enzymes for biocatalysis.

Shinya Fushinobu, Ph.D. 

• Professor
• Head of the Department of Biotechnology
• Graduate School of Agricultural and Life Sciences
• The University of Tokyo
• [email protected]

Biography

Shinya Fushinobu received a Ph.D. degree from the University of Tokyo (1999). He was an assistant professor (1997-2011) and an associate professor (2011-2012) at the same university. He visited Iowa State University in 2006 as a visiting scientist under the supervision of Prof. Peter J. Reilly. His doctoral thesis focuses on the study of the allosteric activation mechanism of bifidobacterial L-lactate dehydrogenase and the X-ray crystallography of an acidophilic xylanase from Aspergillus. He has been engaged in research on the structural and functional analysis of various kinds of enzymes. He was appointed as a full professor in 2012. He published 190 peer-reviewed original papers and received the following awards: JSBBA Award for Encouragement of Young Scientists (2008), Japan Prize in Agricultural Sciences, Achievement Award for Young Scientists (2010), JSAG Distinguished Young Scientist Award (2011), JSPS PRIZE (2016), and JSAG Award (2023). 

Research Interests

• Carbohydrate-Active Enzymes
• Enzymes from gut microbes, especially from bifidobacteria
• Enzymology of various enzymes
• Enzyme classification and nomenclature

Abstract

Structural Analysis of Enzymes Degrading Mycobacterial Lipoarabinomannan and Arabinogalactan

Shinya Fushinobu

Arabinose is notable in that both its L- and D-forms are found in nature. While L-arabinose is abundant in plant cell walls, D-arabinofuranose (D-Araf) is found in certain bacterial cell wall polysaccharides and lipopolysaccharides. Bacteria of the genus Mycobacterium, including Mycobacterium tuberculosis and Mycobacterium leprae, as well as related genera such as Rhodococcus and Nocardia in the Actinomycetales order, are known to contain D-arabinan as components of lipoarabinomannan and arabinogalactan in their cell walls. These mycobacterial D-arabinans are complex polysaccharides composed of α-1,5-linked D-Araf main chains with multiple α-1,3-linked branches, and are capped by β-1,2-D-Araf linkages at their non-reducing termini. The unique glycan structures of mycobacteria contribute to their ability to evade mammalian immune responses.

While L-arabinofuranosidases for degradation of plant polysaccharides have been extensively studied, genes encoding enzymes that act on D-Araf linkages remained unidentified until recently. Our collaborators, Prof. Kiyotaka Fujita and colleagues at Kagoshima University, purified an endo-α-D-arabinanase from the culture supernatant of the soil bacterium Microbacterium arabinogalactanolyticum JCM 9171 and successfully identified its gene.^[1] Within the gene cluster, classified as a polysaccharide utilization locus, three types of D-arabinan-degrading enzymes belonging to glycoside hydrolase (GH) families were identified: two GH183 endo-α-D-arabinanases (EndoMA1 and EndoMA2), a GH172 exo-α-D-arabinofuranosidase (ExoMA1), and a GH116 exo-β-D-arabinofuranosidase (ExoMA2). We have determined the crystal structures of EndoMA1, ExoMA1, and ExoMA2.^[1] In this presentation, I will describe the structural basis for substrate recognition and the reaction mechanisms of these enzymes.

References

[1] M. Shimokawa, A. Ishiwata, T. Kashima, C. Nakashima, J. Li, R. Fukushima, N. Sawai, M. Nakamori, Y. Tanaka, A. Kudo, S. Morikami, N. Iwanaga, G. Akai, N. Shimizu, T. Arakawa, C. Yamada, K. Kitahara, K. Tanaka, Y. Ito, S. Fushinobu, and K. Fujita. Nat. Commun. 14, 5803 (2023)

Romas Kazlauskas, Ph.D. 

• Professor
• Biochemistry, Molecular Biology and Biophysics
• Biotechnology Institute
• University of Minnesota
• [email protected]

Biography

Romas Kazlauskas studied chemistry at the Massachusetts Institute of Technology (Ph.D.) and Harvard University (postdoc with George Whitesides). He worked at General Electric Company (1985-88) and McGill University, Montreal, Canada (1988-2003) and is currently a professor in Biochemistry, Molecular Biology and Biophysics at the University of Minnesota. He has been a visiting professor in Germany, Sweden, South Korea and China. He is an expert in protein engineering of enzymes for biocatalysis.  

Research Interests

• Protein engineering
• Biocatalysis
• Sustainability 

Abstract

Engineering faster enzymes for biocatalysis

Romas Kazlauskas

Engineering faster biocatalytic enzymes remains challenging because residues distant from active sites significantly influence catalysis through poorly understood mechanisms.^[1] While structure-function relationships within active sites are well-characterized, the contribution of remote residues to catalytic efficiency has not been systematically dissected. We sought to identify how distant residues enhance esterase catalysis by engineering a hydroxynitrile lyase (HNL) to acquire esterase activity comparable to the homologous enzyme SABP2. Using computational modeling, X-ray crystallography, and steady-state kinetics, we systematically analyzed how specific distant mutations influence catalysis. Our results reveal three distinct mechanisms by which remote residues enhance esterase activity: (1) preorganization of the catalytic residues through domino-like movements,^[2] (2) creation of a product-release tunnel, and (3) prevention of cosolvent inhibition. Crystallographic analysis showed that distant mutations induced conformational changes extending 10-14 Å from substitution sites. Integrating these insights, we engineered SABP2 variants with 6.5-fold higher catalytic efficiency. These findings provide a framework for rational enzyme engineering that targets allosteric networks rather than active sites alone.

References

[1] R. Buller, S. Lutz, R. J. Kazlauskas, R. Snajdrova, J. C. Moore, U. T. Bornscheuer, Science 2023, 382, eadh8615. https://doi.org/10.1126/science.ahd8615

[2] S. Osuna, WIREs Comput, Mol. Sci. 2021, 11, e1502. https://doi.org/10.1002/wcms.1502