{"@context":{"@vocab":"https://cir.nii.ac.jp/schema/1.0/","rdfs":"http://www.w3.org/2000/01/rdf-schema#","dc":"http://purl.org/dc/elements/1.1/","dcterms":"http://purl.org/dc/terms/","foaf":"http://xmlns.com/foaf/0.1/","prism":"http://prismstandard.org/namespaces/basic/2.0/","cinii":"http://ci.nii.ac.jp/ns/1.0/","datacite":"https://schema.datacite.org/meta/kernel-4/","ndl":"http://ndl.go.jp/dcndl/terms/","jpcoar":"https://github.com/JPCOAR/schema/blob/master/2.0/"},"@id":"https://cir.nii.ac.jp/crid/1362825893553726464.json","@type":"Article","productIdentifier":[{"identifier":{"@type":"DOI","@value":"10.1007/s00253-005-0038-2"}},{"identifier":{"@type":"URI","@value":"http://link.springer.com/content/pdf/10.1007/s00253-005-0038-2.pdf"}},{"identifier":{"@type":"URI","@value":"http://link.springer.com/article/10.1007/s00253-005-0038-2/fulltext.html"}},{"identifier":{"@type":"URI","@value":"http://link.springer.com/content/pdf/10.1007/s00253-005-0038-2"}},{"identifier":{"@type":"PMID","@value":"16012835"}}],"dc:title":[{"@value":"High-level expression and bulk crystallization of recombinant l-methionine γ-lyase, an anticancer agent"}],"description":[{"notation":[{"@value":"L-Methionine gamma-lyase is a pyridoxal 5'-phosphate-dependent enzyme which has tumor selective anticancer activity. An efficient production process for the recombinant enzyme was constructed by using the overexpression plasmid in Escherichia coli, large-scale cultivation, and practical crystallization on an industrial scale. The plasmid was optimized with a promoter and the region of the ribosome-binding site. Plasmid pMGLTrc03, which has a trc promoter and a spacing of 12 nucleotides between the Shine-Dalgarno sequence and the ATG translation initiation codon, was selected as the most suitable plasmid. The transformants produced the enzyme, which intracellularly accumulated at 2.1 mg/ml as an active form and accounted for 43% of the total proteins in the soluble fraction by simple batch fermentation using a 500-l fermentor. The crystals were directly obtained from crude enzyme with 87% yield by a crystallization in the presence of 9.0% polyethylene glycol 6000, 3.6% ammonium sulfate, and 0.18 M sodium chloride using a 100-l crystallizer. After recrystallization, the enzyme was purified by anion-exchange column chromatography to remove endotoxins and by gel filtration for polishing. We prepared 600 g of purified enzyme with a low endotoxin content of sufficient quality for therapeutical use, with a 41% overall yield in the purification process."}]}],"creator":[{"@id":"https://cir.nii.ac.jp/crid/1382825893553726469","@type":"Researcher","foaf:name":[{"@value":"Tomoaki Takakura"}]},{"@id":"https://cir.nii.ac.jp/crid/1382825893553726468","@type":"Researcher","foaf:name":[{"@value":"Takaomi Ito"}]},{"@id":"https://cir.nii.ac.jp/crid/1382825893553726464","@type":"Researcher","foaf:name":[{"@value":"Shigeo Yagi"}]},{"@id":"https://cir.nii.ac.jp/crid/1382825893553726470","@type":"Researcher","foaf:name":[{"@value":"Yoshihide Notsu"}]},{"@id":"https://cir.nii.ac.jp/crid/1382825893553726465","@type":"Researcher","foaf:name":[{"@value":"Takashi Itakura"}]},{"@id":"https://cir.nii.ac.jp/crid/1382825893553726472","@type":"Researcher","foaf:name":[{"@value":"Takumi Nakamura"}]},{"@id":"https://cir.nii.ac.jp/crid/1382825893553726471","@type":"Researcher","foaf:name":[{"@value":"Kenji Inagaki"}]},{"@id":"https://cir.nii.ac.jp/crid/1382825893553726467","@type":"Researcher","foaf:name":[{"@value":"Nobuyoshi Esaki"}]},{"@id":"https://cir.nii.ac.jp/crid/1382825893553726336","@type":"Researcher","foaf:name":[{"@value":"Robert M. Hoffman"}]},{"@id":"https://cir.nii.ac.jp/crid/1382825893553726466","@type":"Researcher","foaf:name":[{"@value":"Akio Takimoto"}]}],"publication":{"publicationIdentifier":[{"@type":"PISSN","@value":"01757598"},{"@type":"EISSN","@value":"14320614"}],"prism:publicationName":[{"@value":"Applied Microbiology and Biotechnology"}],"dc:publisher":[{"@value":"Springer Science and Business Media LLC"}],"prism:publicationDate":"2006-03","prism:volume":"70","prism:number":"2","prism:startingPage":"183","prism:endingPage":"192"},"reviewed":"false","dc:rights":["http://www.springer.com/tdm"],"url":[{"@id":"http://link.springer.com/content/pdf/10.1007/s00253-005-0038-2.pdf"},{"@id":"http://link.springer.com/article/10.1007/s00253-005-0038-2/fulltext.html"},{"@id":"http://link.springer.com/content/pdf/10.1007/s00253-005-0038-2"}],"createdAt":"2005-07-12","modifiedAt":"2024-01-27","foaf:topic":[{"@id":"https://cir.nii.ac.jp/all?q=Antimetabolites,%20Antineoplastic","dc:title":"Antimetabolites, Antineoplastic"},{"@id":"https://cir.nii.ac.jp/all?q=Recombinant%20Proteins","dc:title":"Recombinant Proteins"},{"@id":"https://cir.nii.ac.jp/all?q=Culture%20Media","dc:title":"Culture Media"},{"@id":"https://cir.nii.ac.jp/all?q=Carbon-Sulfur%20Lyases","dc:title":"Carbon-Sulfur Lyases"},{"@id":"https://cir.nii.ac.jp/all?q=Fermentation","dc:title":"Fermentation"},{"@id":"https://cir.nii.ac.jp/all?q=Escherichia%20coli","dc:title":"Escherichia coli"},{"@id":"https://cir.nii.ac.jp/all?q=Crystallization","dc:title":"Crystallization"},{"@id":"https://cir.nii.ac.jp/all?q=Biotechnology","dc:title":"Biotechnology"},{"@id":"https://cir.nii.ac.jp/all?q=Plasmids","dc:title":"Plasmids"}],"relatedProduct":[{"@id":"https://cir.nii.ac.jp/crid/1360568466542206208","@type":"Article","resourceType":"学術雑誌論文(journal article)","relationType":["isReferencedBy"],"jpcoar:relatedTitle":[{"@value":"Application of l-methionine γ-lyase in chiral amino acid analysis"}]},{"@id":"https://cir.nii.ac.jp/crid/1390282681455281920","@type":"Article","resourceType":"学術雑誌論文(journal article)","relationType":["isReferencedBy"],"jpcoar:relatedTitle":[{"@language":"en","@value":"The Role of Amino Acid Residues in the Active Site of <small>L</small>-Methionine γ-lyase from <i>Pseudomonas putida</i>"},{"@value":"The Role of Amino Acid Residues in the Active Site of L-Methionine γ-lyase from Pseudomonas putida"},{"@value":"The Role of Amino Acid Residues in the Active Site of<scp>L</scp>-Methionine γ-lyase from<i>Pseudomonas putida</i>"},{"@value":"The role of amino acid residues in the active site of l-methionine gamma-lyase from Pseudomonas putida"},{"@value":"The role of amino acids residues in the active site of l-methionine γ-lyase from Pseudomonas putida"}]}],"dataSourceIdentifier":[{"@type":"CROSSREF","@value":"10.1007/s00253-005-0038-2"},{"@type":"OPENAIRE","@value":"doi_dedup___::fa716e326c9e35061c07dbd6a2a76600"},{"@type":"CROSSREF","@value":"10.1271/bbb.110906_references_DOI_Ss9hhGaApKHDAJRwhor8zrNPEOt"},{"@type":"CROSSREF","@value":"10.1016/j.ab.2019.05.018_references_DOI_Ss9hhGaApKHDAJRwhor8zrNPEOt"}]}