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Japan Academy Prize to:
Sakayu Shimizu
Emeritus Professor, Kyoto University
Visiting Professor, Toyama Prefectural University
for “Novel Microbial Functions, their Discovery
through Screening, and their Application in theLarge-Scale Production of Useful Compounds”

Outline of the work:
Microorganisms have survived on Earth for nearly four billion years, and their diversity
is immense. Their functions and products are highly varied. Dr. Sakayu Shimizu recognizes
the significance of these microbial communities as crucial resources. He has dedicated yearsof research to exploring the wide-ranging, unknown capabilities of these microorganisms anddelving deep into their discoveries. His perspective revolves around using research findings tobenefit human life by unveiling the untapped potential of these microorganisms.
Dr. Shimizu first took on the challenge of researching lipid production by microorganisms,
choosing arachidonic acid, a physiologically crucial fatty acid for humans, as the target. In
the 1980s, when he began this research, it became clear that animals and plants cannot directly
produce arachidonic acid because certain enzymes are missing the synthesis pathway. It is widelybelieved that microorganisms share the same biosynthetic pathways as plants. He examined
numerous microorganisms for their fatty acid profiles and eventually discovered that the fungus
Mortierella alpina accumulated oil containing arachidonic acid. The yield reached up to 600
mg/g of dried fungal biomass, with approximately 70% of the total fatty acids being arachidonic
acid. This discovery was highly significant as it demonstrated for the first time the existenceof microorganisms capable of directly producing arachidonic acid. Dr. Shimizu subsequently
established a method for the large-scale production of arachidonic acid-containing oil using thisfungus, which is now widely added as an essential component to infant formula, especially forpremature babies.
Dr. Shimizu continued to examine the biosynthetic pathway of polyunsaturated fatty acids(PUFAs) in this fungus. He elucidated various biosynthetic pathways, successfully revealing
them in detail. Furthermore, he achieved individual induction of these pathways, demonstrating
the selective and comprehensive synthesis of 30 rare and valuable PUFAs. During the courseof his research, Dr. Shimizu also discovered the saturation reaction of linoleic acid’s double
bonds by lactic acid bacteria, unraveling the intricacies of this complex process. Additionally, herevealed that sesamin, a component of sesame seeds, acts as a specific inhibitor of the enzyme
in the final stage of arachidonic acid synthesis. Dr. Shimizu’s series of pioneering studies havegiven rise to a new industrial field known as “oil fermentation.”
Dr. Shimizu then embarked on the study of the enzymatic synthesis of optically active

27compounds. Choosing the production process of D-pantothenic acid as his target, he introducedmicrobial reactions into the process of manufacturing its intermediate, D-pantolactone. He discovered a novel enzyme, lactonohydrolase, in a Fusarium fungus. This enzyme selectively hydrolyzed only the D-form of DL-pantolactone, splitting it into D-pantoic acid and L-pantolactone.
The application of this fungus to the optical resolution of DL-pantolactone for the productionof D-pantothenic acid oﬀers a new manufacturing method. This method has been applied on
an industrial scale, producing several thousand tons of D-pantothenic acid annually. The new
method resulted in an approximately 30% reduction in energy consumption and reduced various
environmental burdens compared with conventional chemical optical resolution methods. These
achievements are highly appreciated from the perspective of the current emphasis on sustainable
development goals.
Dr. Shimizu aimed constructed a versatile asymmetric reduction system capable of converting various ketones into optically active alcohols. He organized the gene library of the enzymes
responsible for the asymmetric reduction of various ketones obtained through screening. Then,he designed recombinant Escherichia coli strain that expressed one of these enzymes along withglucose dehydrogenase as an NAD(P)H regenerator. By using this engineered bacterial cell as abiocatalyst, he successfully converted various prochiral ketone compounds into their corresponding optically active alcohols, which had high reaction yields and optical purity. This method has
spread globally as an industrial synthesis approach for diverse optically active intermediates inpharmaceutical synthesis.
Dr. Shimizu’s accomplishments have been documented in approximately 700 research papers. Moreover, he has received several prestigious awards for his work, including from theAmerican Oil Chemists’ Society, the Chemical Society of Japan, the Japan Society for Bioscience, Biotechnology, and Agrochemistry, and the Engineering Foundation, USA, among others. His contributions are highly esteemed both nationally and internationally. In addition, in
2022, he was honored as a Person of Cultural Merit in Japan.
As described above, Dr. Shimizu has made groundbreaking discoveries regarding novel microbial functions through his exploratory research. He has advanced academic studies based onthese findings and has successfully translated the outcomes into high-quality societal implementations, contributing significantly to the development of the biotechnology industry. His researchachievements are highly esteemed, and he is worthy of the Japan Academy Prize in recognitionof his outstanding contributions.

List of Main Publications
1. S. Shimizu, Y. Shinmen, H. Kawashima, K. Akimoto, H. Yamada; Fungal mycelia as anovel source of eicosapentaenoic acid, activation enzyme(s) involved in eicosapentaenoic
acid production at low temperature. Biochem. Biophys. Res. Commun., 150, 335–341(1988).
2. Y. Shinmen, S. Shimizu, K. Akimoto, H. Kawashima, H. Yamada; Production of arachidonic acid by Mortierella fungi: Selection of a potent producer and optimization of culture
conditions for large-scale production. Appl. Microbiol. Biotechnol., 31, 11–16 (1989).

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3. S. Jareonkitmongkol, S. Shimizu, H. Yamada; Occurrence of two non-methylene-interruped
Δ5 polyunsaturated fatty acids in a Δ6-desaturase-defective mutant of the ungus Mortierella
alpina 1S-4. Biochim. Biophys. Acta, 1167, 137–141 (1993).
4. H. Kawashima, K. Akimoto, K. Higashiyama, S. Fujikawa, S. Shimizu; Industrial productionof dihomo-γ-linolenic acid by a Δ5 desaturase-defective mutant of Mortierella alpina 1S-4fungus. J. Am. Oil Chem. Soc., 77, 1135–1138 (2000).
5. E. Sakuradani, M. Kobayashi, S. Shimizu; Δ9-Fatty acid desaturase from arachidonic acidproducing fungus: Unique gene sequence and its heterologous expression in a fungus, Aspergillus. Eur. J. Biochem., 260, 208–216 (1999).
6. E. Sakuradani, M. Kobayashi, T. Ashikari, S. Shimizu; Identification of Δ12-fatty acid desaturase from arachidonic acid-producing Mortierella fungus by heterologous expression inthe yeast Saccharomyces cerevisiae and the fungus, Aspergillus oryzae. Eur. J. Biochem.,
261, 812–820 (1999).
7. S. Takeno, E. Sakuradani, S. Murata, M. Inohara-Ochiai, H. Kawashima, T. Ashikari, S.
Shimizu; Establishment of an overall transformation system for an oil-producing filamentous
fungus, Mortierella alpina 1S-4. Appl. Microbiol. Biotechnol., 65, 419–425 (2004).
8. S. Shimizu, K. Akimoto, Y. Shinmen, H. Kawashima, M. Sugano, H. Yamada; Sesamin isa potent and specific inhibitor of Δ5 desaturase in polyunsaturated fatty acid biosynthesis.
Lipids, 26, 512–516 (1991).
9. J. Ogawa, K. Matsumura, S. Kishino, Y. Omura, S. Shimizu; Conjugated linoleic acid accumulation via 10-hydroxy-12-octadecaenoic acid during microaerobic transformation oflinoleic acid by Lactobacillus acidophilus. Appl. Environ. Microbiol., 67, 1246–1252(2001).
10. S. Kishino, M. Takeuchi, S.B. Park, A. Hirata, N. Kitamura, J. Kunisawa, H. Kiyono, R.
Iwamoto, Y. Isobe, M. Arita, H. Arai, K. Ueda, J. Shima, S. Takahashi, K. Yokozeki, S.
Shimizu, J. Ogawa; Polyunsaturated fatty acid saturation by gut lactic acid bacteria aﬀecting
host lipid composition. Proc. Natl. Acad. Sci. U.S.A., 110, 17808–17813 (2013).
11. S. Shimizu, M. Kataoka, M.C-M. Chung, H. Yamada; Ketopantoic acid reductase of Pseudomonas maltophilia 845: Purification, characterization, and role in pantothenate biosynthesis. J. Biol. Chem., 263, 12077–12084 (1988).
12. S. Shimizu, S. Hattori, H. Hata, H. Yamada; A novel fungal enzyme, NADPH- dependent
carbonyl reductase, showing high specificity to conjugated polyketones: Purification andcharacterization. Eur. J. Biochem., 174, 37–44 (1988).
13. M. Kataoka, S. Shimizu, H. Yamada; Purification and characterization of a novel FMNdependent enzyme, membrane-bound L-(+)-pantoyl lactone dehydrogenase, from Nocardia
asteroids. Eur. J. Biochem., 204, 799–806 (1992).
14. S. Shimizu, M. Kataoka, K. Shimizu, M. Hirakata, K. Sakamoto, H. Yamada; Purificationand characterization of a novel lactonohydrolase, catalyzing the hydrolysis of aldonate lactones and aromatic lactones, from Fusarium oxysporum. Eur. J. Biochem., 209, 383–390(1992).
15. M. Kobayashi, M. Shinohara, C. Sakoh, M. Kataoka, S. Shimizu; Lactone-ring-cleaving
enzyme: Genetic analysis, novel RNA editing, and evolutionary implications. Proc. Natl.

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Acad. Sci. U.S.A., 95, 12787–12792 (1998).
16. M. Kataoka, K. Shimizu, K. Sakamoto, H. Yamada, S. Shimizu; Optical resolution ofracemic pantolactone with a novel fungal enzyme, lactonohydrolase. Appl. Microbiol.
Biotechnol., 43, 974–977 (1995).
17. K. Sakamoto, K. Honda, K. Wada, S. Kita, K. Tsuzaki, H. Nose, M. Kataoka, S. Shimizu;
Practical resolution system for DL-pantoyl lactone using the lactonase from Fusarium oxysporum. J. Biotechnol., 118, 99–106 (2005).
18. M. Kataoka, H. Sakai, T. Morikawa, M. Katoh, T. Miyoshi, S. Shimizu, H. Yamada; Characterization of aldehyde reductase of Sporobolomyces salmonicolor. Biochim. Biophys. Acta,1122, 57–62 (1992).
19. M. Wada, A. Yoshizumi, S. Nakamori, S. Shimizu; Purification and characterization ofmonovalent cation-activated levodione reductase from Corynebacterium aquaticum M-13.
Appl. Environ. Microbiol., 65, 4399–4403 (1999).
20. M. Kataoka, A. Kotaka, A. Hasegawa, M. Wada, A. Yoshizumi, S. Nakamori, S. Shimizu;
Old yellow enzyme from Candida macedoniensis catalyzes stereospecific reduction of C=C
bond of ketoisophorone. Biosci. Biotechnol. Biochem., 66, 2651–2657 (2002).
21. M. Kataoka, K. Yamamoto, H. Kawabata, M. Wada, K. Kita, H. Yanase, S. Shimizu; Stereoselective reduction of ethyl 4-chloro-3-oxobutanoate by Escherichia coli transformant cells
coexpressing the aldehyde reductase and glucose dehydrogenase genes. Appl. Microbiol.
Biotechnol., 51, 486–490 (1999).
22. S. Shimizu, M. Kataoka, M. Katoh, T. Morikawa, T. Miyoshi, H. Yamada; Stereoselective
reduction of ethyl 4-chloro-3-oxobutanoate by a microbial aldehyde reductase in an organic
solvent-water diphasic system. Appl. Environ. Microbiol., 56, 2374–2377 (1990).
23. N. Kizaki, Y. Yasohara, J. Hasegawa, M. Wada, M. Kataoka, S. Shimizu; Synthesis of optically pure ethyl (S)-4-chloro-3-hydroxybutanoate by Escherichia coli transformant cells
coexpressing the carbonyl reductase and glucose dehydrogenase genes. Appl. Microbiol.
Biotechnol., 55, 590–595 (2001).
24. M. Wada, A. Yoshizumi, Y. Noda, M. Kataoka, S. Shimizu, H. Takagi, S. Nakamori; Production of a doubly chiral compound, (4R,6R)-4-hydroxy-2,2,6- trimethyl cyclohexanone, bytwo-step enzymatic asymmetric reduction. Appl. Environ. Microbiol., 69, 933–937 (2003).