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Takeji Shibatani

Bio: Takeji Shibatani is an academic researcher. The author has contributed to research in topics: Serratia marcescens & Lipase. The author has an hindex of 12, co-authored 35 publications receiving 404 citations.

Papers
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Journal ArticleDOI
TL;DR: The Serratia marcescens S‐layer protein is secreted to the cultured media in Escherichia coli cells carrying the Lip exporter, indicating that the S. marces censored protein is strictly recognized by the Lip system, the first report concerning secretion of an S-layer protein via its own secretion system.
Abstract: The Serratia marcescens Lip exporter belonging to the ATP-binding cassette (ABC) exporter is known to be involved in signal peptide-independent extracellular secretion of a lipase and a metalloprotease. Although the genes of secretory proteins and their ABC exporters are usually all reported to be linked in several gram-negative bacteria, neither the lipase nor the protease gene is located close to the Lip exporter genes, lipBCD. A gene (slaA) located upstream of the lipBCD genes was cloned, revealing that it encodes a polypeptide of 100 kDa and is partially similar to the Caulobacter crescentus paracrystalline cell surface layer (S-layer) protein. The Lip exporter-deficient mutants of S. marcescens failed to secrete the SlaA protein. Electron micrography demonstrated the cell surface layer of S. marcescens. The S-layer protein was secreted to the cultured media in Escherichia coli cells carrying the Lip exporter. Three ABC exporters, Prt, Has and Hly systems, could not allow the S-layer secretion, indicating that the S. marcescens S-layer protein is strictly recognized by the Lip system. This is the first report concerning secretion of an S-layer protein via its own secretion system.

90 citations

Journal ArticleDOI
TL;DR: In this article, an asymmetric (±)-trans -3-(4-Methoxyphenyl) glycidic acid methyl ester (MPGM) was hydrolyzed to (+)-(2 S, 3 R )-3-(methoxymhenyl)-glycidric acid and methanol using a hollow-fiber ultrafiltration membrane.

57 citations

Journal ArticleDOI
TL;DR: A lipase from Serratia marcescens was selected as an asymmetric hydrolytic enzyme for trans-3-(4-methoxyphenyl)glycidic acid methyl ester [(±)-MPGM], a key intermediate in the synthesis of diltiazem hydrochloride that is useful as a coronary vasodilator.
Abstract: A lipase from Serratia marcescens was selected as an asymmetric hydrolytic enzyme for trans-3-(4-methoxyphenyl)glycidic acid methyl ester [(±)-MPGM], a key intermediate in the synthesis of diltiazem hydrochloride that is useful as a coronary vasodilator. This lipase has high enantioselectivity (E=135) and was applied to the industrial production of the optically active intermediate of diltiazem using two-phase reaction system of organic solvent–water. Introduction of enzymatic reaction into the chemical synthetic route of diltiazem reduces the number of processes from nine to five. Analyses of the secretion mechanism of the lipase from S. marcescens cell membrane revealed that lipase (LipA), metalloprotease (PrtA), cell surface protein (SlaA) and flagellin are secreted via ABC-transporter, which is a common secreting mechanism in Gram-negative bacteria other than N-terminal signal peptide-dependent secreting mechanism. Molecular cloning of both the lipA gene, which codes the lipase protein, and lipBCD genes, which code the secretion device proteins, enable the production of the lipase by the self-cloning strain 140-fold as compared to the wild type strain. Immobilization of the lipase on a hollow fiber type membrane reactor contributes to the repeated use of enzyme and to efficient separation of the reaction product. Thus, enzymatic reaction and product separation are achieved simultaneously.

36 citations

Journal ArticleDOI
TL;DR: An engineering analysis of l-lysine racemization and microbial degradation was carried out to establish the basis of process design for d-lysines production.
Abstract: In order to develop a practical process for D-lysine production from L-lysine, successive chemical racemization and microbial asymmetric degradation were investigated. The racemization of L-lysine proceeded quantitatively at elevated temperatures. A sample of 1000 strains of bacteria, fungi, yeast and actinomyces were screened for the ability to degrade L-lysine asymmetrically. Microorganisms belonging to the Achromobacter, Agrobacterium, Candida, Comamonas, Flavobacterium, Proteus, Providencia, Pseudomonas and Yarrowia genera exhibited a high L-lysine-degrading activity. Comamonas testosteroni IAM 1048 was determined to be the best strain and used as a biocatalyst for eliminating the L isomer. The degradation rate of L-lysine with C. testosteroni IAM 1048 was influenced by pH, temperature and agitation speed. Under the optimal conditions, the L isomer in a 100-g/l mixture of racemic lysine was completely degraded within 72 h, with 47 g D-lysine/l left in the reaction mixture. Crystalline D-lysine, with a chemical purity greater than 99% and optical purity of 99.9% enantiomeric excess, was obtained at a yield of 38% from the reaction mixture by simple purification. An engineering analysis of L-lysine racemization and microbial degradation was carried out to establish the basis of process design for D-lysine production.

19 citations


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Journal ArticleDOI
TL;DR: Novel biotechnological applications have been successfully established using lipases for the synthesis of biopolymers and biodiesel, the production of enantiopure pharmaceuticals, agrochemicals, and flavour compounds.

1,315 citations

Journal ArticleDOI
TL;DR: The latest trend in lipase research is the development of novel and improved lipases through molecular approaches such as directed evolution and exploring natural communities by the metagenomic approach.
Abstract: Lipases, triacylglycerol hydrolases, are an important group of biotechnologically relevant enzymes and they find immense applications in food, dairy, detergent and pharmaceutical industries. Lipases are by and large produced from microbes and specifically bacterial lipases play a vital role in commercial ventures. Some important lipase-producing bacterial genera include Bacillus, Pseudomonas and Burkholderia. Lipases are generally produced on lipidic carbon, such as oils, fatty acids, glycerol or tweens in the presence of an organic nitrogen source. Bacterial lipases are mostly extracellular and are produced by submerged fermentation. The enzyme is most commonly purified by hydrophobic interaction chromatography, in addition to some modern approaches such as reverse micellar and aqueous two-phase systems. Most lipases can act in a wide range of pH and temperature, though alkaline bacterial lipases are more common. Lipases are serine hydrolases and have high stability in organic solvents. Besides these, some lipases exhibit chemo-, regio- and enantioselectivity. The latest trend in lipase research is the development of novel and improved lipases through molecular approaches such as directed evolution and exploring natural communities by the metagenomic approach.

1,077 citations

Journal ArticleDOI
TL;DR: Three-dimensional structures of bacterial lipases were solved to understand the catalytic mechanism of lipase reactions and will enable researchers to tailor new lipases for biotechnological applications.
Abstract: ▪ Abstract Bacteria produce and secrete lipases, which can catalyze both the hydrolysis and the synthesis of long-chain acylglycerols. These reactions usually proceed with high regioselectivity and enantioselectivity, and, therefore, lipases have become very important stereoselective biocatalysts used in organic chemistry. High-level production of these biocatalysts requires the understanding of the mechanisms underlying gene expression, folding, and secretion. Transcription of lipase genes may be regulated by quorum sensing and two-component systems; secretion can proceed either via the Sec-dependent general secretory pathway or via ABC transporters. In addition, some lipases need folding catalysts such as the lipase-specific foldases and disulfide-bond–forming proteins to achieve a secretion-competent conformation. Three-dimensional structures of bacterial lipases were solved to understand the catalytic mechanism of lipase reactions. Structural characteristics include an α/β hydrolase fold, a catalytic ...

1,072 citations

Journal ArticleDOI
TL;DR: Cell walls are an important structural component of prokaryotic organisms and essential for many aspects of their life, and the diverse structures of the outermost boundary layers strongly reflect adaptations to specific ecological and environmental conditions.
Abstract: Cell walls are an important structural component of prokaryotic organisms and essential for many aspects of their life. Particularly, the diverse structures of the outermost boundary layers strongly reflect adaptations of organisms to specific ecological and environmental conditions ([6][1]). Over

757 citations