OKANO, Kenji |
|---|
Faculty, Department/Institute
- Faculty of Chemistry, Materials and Bioengineering Department of Life Science and Biotechnology
Academic status (qualification)
- Associate Professor Apr. 1,2022
Undergraduate Degrees・University
- Kobe University Faculty of Engineering2004
- Kobe University Graduate School of Science and Technology2009
- Kobe University Graduate School of Science and Technology2006
Academic Degrees
- 博士（工学） 神戸大学
Awards
- Apr. 2022
- 第73回日本生物工学会大会「トピックス賞」 Oct. 2021
- 第29回生物工学論文賞 Oct. 2021
- Oct. 2015
- Best Poster Award Nov. 2014(Young Asian Biological Engineers' Community)
Academic Associations
| 所属学会・団体名 | 役職名 (役職在任期間) |
|---|---|
| The Society for Biotechnology, Japan | |
| THE SOCIETY OF CHEMICAL ENGINEERS, JAPAN | |
| Japanese Society of Enzyme Engineering | |
| JAPAN SOCIETY FOR BIOSCIENCE, BIOTECHNOLOGY, AND AGROCHEMISTRY | |
| JAPAN SOCIETY FOR LACTIC ACID BACTERIA |
Intellectual Property Rights
- Recycling method of S-adenosylmethionine application number：特願2018-024361 (Feb. 14,2018)
- Homolactic Fermentation from Pentose application number：特願JP2010052216 (Feb. 15,2010)
- application number：特開WO2010095600 (Aug. 26,2010)
Research Publications
PapersIn refereed2022/4～10.1002/biot.202100331
PapersIn refereed2021/12～10.1016/j.jbiosc.2021.09.003
PapersIn refereed2021/5/19～10.1016/j.jbiosc.2021.04.009
PapersIn refereed2021/5/14～10.1128/AEM.00541-21
PapersIn refereed2021～10.3389/fbioe.2021.661096
PapersIn refereed2021～
PapersIn refereed2020/9/2～10.3390/catal10091001
PapersIn vitro reconstitution of non-phosphorylative Entner-Doudoroff pathway for lactate production.In refereedKenji Okano, Qianqin Zhu, Kohsuke HondaJournal of Bioscience and Bioengineering129,3,269-2752020/3～10.1016/j.jbiosc.2019.09.010In vitro metabolic engineering is an emerging framework for bioproduction systems, in which synthetic metabolic pathways are constructed using a limited number of enzymes. Employment of thermophilic enzymes as catalytic elements in pathways enables the use of simple heat purification of recombinantly expressed enzymes. However, thermophilic enzymes are generally incompatible with thermo-labile substrates and intermediates. In previous work, we showed that lactate production through a non-ATP forming chimeric Embden-Meyerhof (EM) pathway required careful adjustment of the metabolic fluxes by continuous substrate feeding and optimization of enzyme ratios to prevent the accumulation and degradation of thermo-labile intermediates (Ye et al., Microb. Cell Fact., 11, 120, 2012). In the study reported here, we constructed an in vitro non-phosphorylative Entner-Doudoroff (np-ED) pathway. Because of the high thermal stability of the metabolic intermediates in the np-ED pathway, it could prevent degradation of accumulated metabolic intermediates caused by inconstant metabolic fluxes, and batch-mode production of lactate in which the concentrations of the substrate and metabolic intermediates change dynamically could be achieved. By combining the enzymes involved in the np-ED pathway and lactate dehydrogenase, 20.9 mM lactate was produced from 10 mM glucose and 1 mM gluconate in 6 h.
PapersIn refereed2020～10.1264/jsme2.ME20074
PapersIn vitro production of cysteine from glucose.In refereedHanatani Y, Imura M, Taniguchi H, Okano K, Toya Y, Iwakiri R, Honda KApplied Microbiology and Biotechnology103,19,8009-80192019/10～10.1007/s00253-019-10061-40175-7598Cysteine is a commercially valuable amino acid with an increasing demand in the food, cosmetic, and pharmaceutical industries. Although cysteine is conventionally manufactured by extraction from animal proteins, this method has several problems, such as troublesome waste-water treatment and incompatibility with some dietary restrictions. Fermentative production of cysteine from plant-derived substrates is a promising alternative for the industrial production of cysteine. However, it often suffers from low product yield as living organisms are equipped with various regulatory systems to control the intracellular cysteine concentration at a moderate level. In this study, we constructed an in vitro cysteine biosynthetic pathway by assembling 11 thermophilic enzymes. The in vitro pathway was designed to be insensitive to the feedback regulation by cysteine and to balance the intra-pathway consumption and regeneration of cofactors. A kinetic model for the in vitro pathway was built using rate equations of individual enzymes and used to optimize the loading ratio of each enzyme. Consequently, 10.5 mM cysteine could be produced from 20 mM glucose through the optimized pathway. However, the observed yield and production rate of the assay were considerably lower than those predicted by the model. Determination of cofactor concentrations in the reaction mixture indicated that the inconsistency between the model and experimental assay could be attributed to the depletion of ATP and ADP, likely due to host-derived, thermo-stable enzyme(s). Based on these observations, possible approaches to improve the feasibility of cysteine production through an in vitro pathway have been discussed.
PapersDeveloping a single strain for in vitro salvage synthesis of NAD+ at high temperatures and its potential for bioconversion.In refereedTaniguchi H, Imura M, Okano K, Honda KMicrobial Cell Factories18,1,75-2019/4～10.1186/s12934-019-1125-x
PapersExpression of engineered carbonyl reductase from Ogataea minuta in Rhodococcus opacus and its application to whole-cell bioconversion in anhydrous solvents.In refereedHonda K, Ono T, Okano K, Miyake R, Dekishima Y, Kawabata HJournal of Bioscience and Bioengineering127,2,145-1492019/2～10.1016/j.jbiosc.2018.07.0111389-1723
PapersThe directions and methods of genetic engineering and cultivation of lactic acid bacteria for the beginnersIn refereed30,1,8-172019～1343-327X
PapersDe novo design of biosynthetic pathways for bacterial production of bulk chemicals and biofuels.In refereedOkano K, Honda K, Taniguchi H, Kondo AFEMS Microbiology Letters365,20,fny215-2018/10～10.1093/femsle/fny2150378-1097
Book2018/8～486043563X
BookPhosphorus recovery and recyclingOkano, K, Ohtake, H, Kunisada, H, Takano, H, Toda, MSpringer Nature Singapore5262018/6～9789811080302
PapersIn refereed2018/5/1～10.1002/biot.2017005171860-7314
Book2018/3～4781313175
PapersIn refereed2018/1～10.1007/s13280-017-0976-90044-7447
Book2017/11～4254141041
Keynote addressIn vitro metabolic engineering for the salvage synthesis of NAD+Kenji Okano, Kohsuke HondaYABEC20172017/10/19～Xi’an, China
Keynote addressProduction of optically pure D-lactic acid from renewable resourcesKenji OkanoThe 8th China-Japan symposium on chemical engineering2017/10/15～Beijing, China
PapersIn vitro bioconversion of chitin to pyruvate with thermophilic enzymesIn refereedKohsuke Honda, Keisuke Kimura, Pham Huynh Ninh, Hironori Taniguchi, Kenji Okano, Hisao OhtakeJOURNAL OF BIOSCIENCE AND BIOENGINEERING124,3,296-3012017/9～10.1016/j.jbiosc.2017.04.0131389-1723Chitin is the second most abundant organic compound on the planet and thus has been regarded as an alternative resource to petroleum feedstocks. One of the key challenges in the biological conversion of biomass-derived polysaccharides, such as cellulose and chitin, is to close the gap between optimum temperatures for enzymatic saccharification and microbial fermentation and to implement them in a single bioreactor. To address this issue, in the present study, we aimed to perform an in vitro, one-pot bioconversion of chitin to pyruvate, which is a precursor of a wide range of useful metabolites. Twelve thermophilic enzymes, including that for NAD(+) regeneration, were heterologously produced in Escherichia coli and semi-purified by heat treatment of the crude extract of recombinant cells. When the experimentally decided concentrations of enzymes were incubated with 0.5 mg mL(-1) colloidal chitin (equivalent to 2.5 mM N-acetylglucosamine unit) and an adequate set of cofactors at 70 degrees C, 0.62 mM pyruvate was produced in 5 h. Despite the use of a cofactor-balanced pathway, determination of the pool sizes of cofactors showed a rapid decrease in ATP concentration, most probably due to the thermally stable ATP-degrading enzyme(s) derived from the host cell. Integration of an additional enzyme set of thermophilic adenylate kinase and polyphosphate kinase led to the deceleration of ATP degradation, and the final product titer was improved to 2.1 mM. (C) 2017, The Society for Biotechnology, Japan. All rights reserved.
PapersIn refereed2017/9～10.1128/JB.00359-170021-9193
Keynote addressProduction of optically pure D-lactic acid from renewable resourcesKenji Okano, Shinji Hama, Tsutomu Tanaka, Hideo Noda, Akihiko Kondo, Kohsuke Honda12th International Symposium on Lactic Acid Bacteria2017/8/29～Egmond aan Zee, Netherland
PapersModules for in vitro metabolic engineering: Pathway assembly for bio-based production of value-added chemicalsIn refereedHironori Taniguchi, Kenji Okano, Kohsuke HondaSynthetic and Systems Biotechnology2,2,65-742017/6/1～10.1016/j.synbio.2017.06.0022405-805XBio-based chemical production has drawn attention regarding the realization of a sustainable society. In vitro metabolic engineering is one of the methods used for the bio-based production of value-added chemicals. This method involves the reconstitution of natural or artificial metabolic pathways by assembling purified/semi-purified enzymes in vitro. Enzymes from distinct sources can be combined to construct desired reaction cascades with fewer biological constraints in one vessel, enabling easier pathway design with high modularity. Multiple modules have been designed, built, tested, and improved by different groups for different purpose. In this review, we focus on these in vitro metabolic engineering modules, especially focusing on the carbon metabolism, and present an overview of input modules, output modules, and other modules related to cofactor management.
PapersIn refereed2017/6～10.1016/j.jbiosc.2017.01.0161389-1723
PapersIn vitro bioconversion of chitin to pyruvate with thermophilic enzymes. J. Biosci. Bioeng.In refereedHonda K, Kimura K, Ninh PH, Taniguchi H, Okano K, Ohtake HJ. Biosci. Bioeng.,,-2017/5～
PapersIn refereed2017/3～10.1007/s00253-016-7976-80175-7598
PapersMetabolic engineering of lactic acid bacteria for production of D-lactic acid from unutilized biomass resources岡野 憲司, 濵 真司, 田中 勉, 野田 秀夫, 近藤 昭彦JATAFFジャーナル = JATAFF journal : 農林水産技術5,3,33-372017/3～2187-4948
Keynote addressProduction of optically pure D-lactic acid from renewable resourcesKenji Okano, Shinji Hama, Tsutomu Tanaka, Chiaki Noda, Hideo Noda, Akihiko KondoYABEC 20162016/10/28～Miyazaki
PapersIn refereed2016/10～10.1016/j.seppur.2016.06.0401383-5866
PapersIn refereed2016/6～10.1016/j.seppur.2016.03.0541383-5866
Keynote address2016/5/18～
PapersIn refereed2016/5～10.1016/j.ymben.2016.02.0051096-7176
Papers2016/4～0387-1037
PapersIn refereed2016/1～10.1371/journal.pone.01461461932-6203
PapersDirected evolution of thermotolerant malic enzyme for improved malate productionYe Xiaothing生物工学会誌 : seibutsu-kogaku kaishi94,2,-2016～
Papers1-Butanol production by in vitro metabolic engineeringOKANO Kenji, HONDA Kohsukeバイオサイエンスとインダストリー = Bioscience & industry73,6,481-4822015/11～0914-8981
Keynote addressAssembly and multiple gene expression of thermophilic enzymes in Escherichia coli for in vitro metabolic engineeringOkano K, Honda K, Ohtake HYABEC 20152015/10/15～Chuncheon, Korea
PapersDevelopment and implementation of technologies for recycling phosphorus in secondary resources in JapanIn refereedHisao Ohtake, Kenji OkanoGlobal Environmental Research19,1,49-652015/5～
PapersIn refereed2015/4～10.1016/j.seppur.2015.01.0431383-5866
Keynote addressAssembly and multiple gene expression of thermophilic enzymes in Escherichia coli for in vitro metabolic engineeringOkano K, Honda K, Ohtake HBiotechnology and Chemistry for Green Growth2015/3/10～
PapersIn refereed2015/1～10.1002/bit.253380006-3592
Keynote addressIn vitro metabolic engineering employing thermophilic enzymes –a novel, simple technology for designing a chimeric metabolic pathwayKenji Okano, Kohsuke Honda, Hisao OhtakeYABEC20142014/11/7～Chiayi, Taiwan
PapersIn vitro conversion of glycerol to lactate with thermophilic enzymesIn refereedJaturapaktrarak C, Napathorn SC, Cheng M, Okano K, Ohtake H, Honda KBioresources and Bioprocessing1,18,-2014/10～
PapersIn refereed2014/7～10.1007/s00253-014-5668-90175-7598
PapersIn refereed2014/4/28～10.1002/9781118845394.ch13
BookBioprocessing of Renewable Resources to Commodity BioproductsOkano, K, Tanaka, T, Kondo AWilley5842014/4～9781118175835
PapersIn refereed2014/2～10.1016/j.jbiosc.2013.07.0051389-1723
Papers1P-153 Succinate production by in vitro metabolic engineeringShimada Daiki, Hashimoto Takahiro, Okano Kenji, Honda Kohsuke, Ohtake Hisao66,,56-562014～
Papers1P-151 Construction of non-phosphorylated Entner-Doudoroff pathway by in vitro metabolic engineering and its application to lactate productionOkano Kenji, Zhu Qianqin, Honda Kohsuke, Ohtake Hisao66,,55-552014～
Papers1P-152 Multiple-gene-expression of thermophilic enzymes for one-step construction of in vitro metabolic pathway :Xiaoyu Bei, Ninh Huynh Pham, Kosuke Kosuke, Kenji Okano, Hisao Ohtake66,,56-562014～
PapersInnovative P Recovery Process Using Amorphous Calcium Silicate Hydrates岡野 憲司, 國貞 眞司, 高野 博幸ファインケミカル : 調査・資料・報道・抄録42,12,24-292013/12～0913-6150
PapersIn refereed2013/10～10.1186/1475-2859-12-911475-2859
PapersIn refereed2013/5～10.1016/j.watres.2013.01.0520043-1354
PapersIn refereed2013/3～10.1016/j.jbiotec.2012.11.0110168-1656
PapersIn refereed2013/3～10.1128/AEM.03752-120099-2240
PapersIn refereed2013～10.1016/j.ymben.2013.09.0061096-7184
Papers3P-162 Construction of a shortcut lactate production pathway by synthetic metabolic engineering :Zhu Qianqin, Okano Kenji, Honda Kohsuke, Ohtake Hisao65,,228-2282013～
Keynote addressNovel technology for phosphorus recycling using amorphous calcium silicate hydrates (A-CSHs)Kenji Okano, Kohsuke Honda, Hisao OhtakeEcobalance 20122012/11/21～Keio University, Kanagawa
Keynote addressNovel technology for phosphorus recycling using amorphous calcium silicate hydrates (A-CSHs)Kenji Okano, Kohsuke Honda, Hisao OhtakeYABEC 20122012/10/27～Tokushima University, Tokushima
Papers2012/10/25～0919-3758
PapersIdentification of the Replication Region of a 111-kb Circular Plasmid from Rhodococcus opacus B-4 by lambda Red Recombination-Based Deletion AnalysisIn refereedKohsuke Honda, Makoto Imura, Kenji Okano, Takeshi Omasa, Junichi Kato, Hisao OhtakeBIOSCIENCE BIOTECHNOLOGY AND BIOCHEMISTRY76,9,1758-17642012/9～10.1271/bbb.1203620916-8451The replication region of the 111-kb circular plasmid pKNR from Rhodococcus opacus B-4 was identified. A PCR-based deletion analysis using the lambda Red recombination technique followed by restriction digestion and PCR-amplification analyses revealed that a 2.5-kb fragment covering one putative open reading frame (ORF) was involved in the replication of pKNR. The product of this ORF showed significant similarity to a functionally unknown protein encoded in the replication region of the 70-kb circular plasmid of Clavibacter michiganensis and to ones in other bacterial large circular plasmids. These observations suggest that the product of the identified ORF and its orthologs can serve as novel replication proteins for large circular bacterial plasmids.
PapersIn refereed2012/9～10.1186/1475-2859-11-1201475-2859
Papers2012/8/1～0453-073X
Papers2012/6/25～0919-3758
Keynote address2012/6/22～
Book2012/4～4781305636
PapersIn refereed2012/4～10.1016/j.jbiosc.2011.11.0161389-1723
Papers4Ap08 Synthetic metabolic engineering : A novel, simple technology for designing a chimeric metabolic pathway(SBJ/JST Joint Symposium IV-System & Synthetic Biotechnology-) :HONDA Kohsuke, YE Xiaoting, OKANO Kenji, OHTAKE Hisao64,,77-772012～
Papers4Ha06 Alteration of the coenzyme specificity of the malic enzyme from Thermococcus kodakarensis KOD1 and its utilization in synthetic metabolic pathwayMorimoto Yumi, YE Xiaoting, Honda Kohsuke, Okano Kenji, Ohtake Hisao64,,230-2302012～
Papers4Ha07 Use of lipophilic bacterium Rhodococcus opacus B4 in organic solvents as whole-cell catalystWada Mayumi, Honda Kohsuke, Okano Kenji, Ohtake Hisao64,,231-2312012～
Papers4Ha09 Construction of the artificial pyruvate oxidation pathway by synthetic metabolic engineeringImagawa Takashi, Honda Kousuke, Okano Kenji, Ohtake Hisao64,,231-2312012～
Papers2Bp02 Biocatalytic activity of heat-treated cells of the hydrophobic bacterium Rhodococcus rhodochrous NBRC15564 in essentially wate-free environmentsHibino Aiko, Honda Kousuke, Okano Kenji, Ohtake Hisao64,,28-282012～
PapersIn refereed2011/11～10.1016/j.bej.2011.08.0021369-703X
PapersIn refereed2011/10～10.1007/s00253-011-3356-60175-7598
PapersIn refereed2011/9～10.1007/s00253-011-3342-z0175-7598
PapersD-lactic acid production from cellooligosaccharides and beta-glucan using L-LDH gene-deficient and endoglucanase-secreting Lactobacillus plantarumIn refereedKenji Okano, Qiao Zhang, Shogo Yoshida, Tsutomu Tanaka, Chiaki Ogino, Hideki Fukuda, Akihiko KondoAPPLIED MICROBIOLOGY AND BIOTECHNOLOGY85,3,643-6502010/1～10.1007/s00253-009-2111-80175-7598In order to achieve direct fermentation of an optically pure d-lactic acid from cellulosic materials, an endoglucanase from a Clostridium thermocellum (CelA)-secreting plasmid was introduced into an l-lactate dehydrogenase gene (ldhL1)-deficient Lactobacillus plantarum (a dagger ldhL1) bacterial strain. CelA expression and its degradation of beta-glucan was confirmed by western blot analysis and enzyme assay, respectively. Although the CelA-secreting a dagger ldhL1 assimilated cellooligosaccharides up to cellohexaose (although not cellotetraose), the main end product was acetic acid, not lactic acid, due to the conversion of lactic acid to acetic acid. Cultivation under anaerobic conditions partially suppressed this conversion resulting in the production of 1.27 g/l of D-lactic acid with a high optical purity of 99.5% from a medium containing 2 g/l of cellohexaose. Subsequently, D-lactic acid fermentation from barley beta-glucan was carried out with the addition of Aspergillus aculeatus beta-glucosidase produced by recombinant Aspergillus oryzae and 1.47 g/l of D-lactic was produced with a high optical purity of 99.7%. This is the first report of direct lactic acid fermentation from beta-glucan and a cellooligosaccharide that is a more highly polymerized sugar than cellotriose.
PapersIn refereed2010/1～10.1007/s00253-009-2280-50175-7598
PapersIn refereed2009/12～10.1128/AEM.01692-090099-2240
PapersIn refereed2009/8～10.1128/AEM.00573-090099-2240
Papers2009/3～0387-1037
PapersEfficient Production of Optically Pure D-Lactic Acid from Raw Corn Starch by Using a Genetically Modified L-Lactate Dehydrogenase Gene-Deficient and alpha-Amylase-Secreting Lactobacillus plantarum StrainIn refereedKenji Okano, Qiao Zhang, Satoru Shinkawa, Shogo Yoshida, Tsutomu Tanaka, Hideki Fukuda, Akihiko KondoAPPLIED AND ENVIRONMENTAL MICROBIOLOGY75,2,462-4672009/1～10.1128/AEM.01514-080099-2240In order to achieve direct and efficient fermentation of optically pure D-lactic acid from raw corn starch, we constructed L-lactate dehydrogenase gene (ldhL1)-deficient Lactobacillus plantarum and introduced a plasmid encoding Streptococcus bovis 148 alpha-amylase (AmyA). The resulting strain produced only D-lactic acid from glucose and successfully expressed amyA. With the aid of secreting AmyA, direct D-lactic acid fermentation from raw corn starch was accomplished. After 48 h of fermentation, 73.2 g/liter of lactic acid was produced with a high yield (0.85 g per g of consumed sugar) and an optical purity of 99.6%. Moreover, a strain replacing the ldhL1 gene with an amyA-secreting expression cassette was constructed. Using this strain, direct D-lactic acid fermentation from raw corn starch was accomplished in the absence of selective pressure by antibiotics. This is the first report of direct D-lactic acid fermentation from raw starch.
Keynote addressA. Novel cell surface display on lactic acid bacteria and its application to lactic acid production from starchy materialsKenji Okano, Qiao Zhang, Tsutomu Tanaka, Hideki Fukuda, Akihiko Kondo2008 AIChE Annual Meeting2008/11/20～Philadelphia, USA
Papers2008/11/1～
PapersIn refereed2008/2～10.1128/AEM.02012-070099-2240
Keynote addressUseful compound production from biomass resources using cell surface engineered microorganismsKenji Okano, Tsutomu Tanaka, Chiaki Ogino, Hideki Fukuda, Akihiko Kondo2007/12/5～Osaka
PapersImprovement in lactic acid production from starch using alpha-amylase-secreting Lactococcus lactis cells adapted to maltose or starchIn refereedKenji Okano, Sakurako Kimura, Junya Narita, Hideki Fukuda, Akihiko KondoAPPLIED MICROBIOLOGY AND BIOTECHNOLOGY75,5,1007-10132007/7～10.1007/s00253-007-0905-00175-7598To achieve direct and efficient lactic acid production from starch, a genetically modified Lactococcus lactis IL 1403 secreting alpha-amylase, which was obtained from Streptococcus bovis 148, was constructed. Using this strain, the fermentation of soluble starch was achieved, although its rate was far from efficient (0.09 g l(-1) h(-1) lactate). High-performance liquid chromatography revealed that maltose accumulated during fermentation, and this was thought to lead to inefficient fermentation. To accelerate maltose consumption, starch fermentation was examined using L. lactis cells adapted to maltose instead of glucose. This led to a decrease in the amount of maltose accumulation in the culture, and, as a result, a more rapid fermentation was accomplished (1.31 g l(-1) h(-1) lactate). Maximum volumetric lactate productivity was further increased (1.57 g l(-1) h(-1) lactate) using cells adapted to starch, and a high yield of lactate (0.89 g of lactate per gram of consumed sugar) of high optical purity (99.2% of L-lactate) was achieved. In this study, we propose a new approach to lactate production by alpha-amylase-secreting L. lactis that allows efficient fermentation from starch using cells adapted to maltose or starch before fermentation.
Book2007/3～4860431065
PapersIn refereed2006/11～10.1007/s00253-006-0477-40175-7598
Keynote addressEfficient lactic acid production from starch by maltose- or starch-adapted α-amylase-secreting Lactococcus lactisKenji Okano, Sakurako Kimura, Hideki Fukuda, Akihiko KondoYABEC 20062006/10/28～Kaohsiung, Taiwan
Papers2006/9～0919-3758
Papers2006/7～1342-3037
PapersIn refereed2006/5～10.1007/s00253-005-0111-x0175-7598
PapersDisplay of alpha-amylase on the surface of Lactobacillus casei cells by use of the PgsA anchor protein, and production of lactic acid from starchIn refereedJ Narita, K Okano, T Kitao, S Ishida, T Sewaki, MH Sung, H Fukuda, A KondoAPPLIED AND ENVIRONMENTAL MICROBIOLOGY72,1,269-2752006/1～10.1128/AEM.72.1.269-275.20060099-2240We developed a new cell surface engineering system based on the PgsA anchor protein from Bacillus subtilis. In this system, the N terminus of the target protein was fused to the PgsA protein and the resulting fusion protein was expressed on the cell surface. Using this new system, we constructed a novel starch-degrading strain of Lactobacillus casei by genetically displaying a-amylase from the Streptococcus bovis strain 148 with a FLAG peptide tag (AmyAF). Localization of the PgsA-AmyA-FLAG fusion protein on the cell surface was confirmed by immunofluorescence microscopy and flow cytometric analysis. The lactic acid bacteria which displayed AmyAF showed significantly elevated hydrolytic activity toward soluble starch. By fermentation using AmyAF-displaying L. casei cells, 50 g/liter of soluble starch was reduced to 13.7 g/liter, and 21.8 g/liter of lactic acid was produced within about 24 h. The yield in terms of grams of lactic acid produced per gram of carbohydrate utilized was 0.60 g per g of carbohydrate consumed at 24 h. Since AmyA was immobilized on the cells, cells were recovered after fermentation and used repeatedly. During repeated utilization of cells, the lactic acid yield was improved to 0.81 g per g of carbohydrate consumed at 72 h. These results indicate that efficient simultaneous saccharification and fermentation from soluble starch to lactic acid were carried out by recombinant L. casei cells with cell surface display of AmyA.
Papers2005～10.11491/scej.2005.0.255.0
Courses Taught
- Experiments of Chemistry
- Orientation Seminar
- Microbiology III
- Experiments of Biology
- Experiments of Biotechnology
- Microbiology IV
- Exercises in Thesis Projects I and II
- Thesis Projects I
- Thesis Projects II
- Science and Technology English II
- Advanced Molecular Microbiology
- SeminarI（Biotechnology)
- SeminarII（Biotechnology)
- Advanced Molecular Microbiology
- Personal Information
- Research Activities
- Research Activities
- Community Service
- Courses Taught