Chemistry: molecular biology and microbiology – Micro-organism – tissue cell culture or enzyme using process... – Preparing nitrogen-containing organic compound
Reexamination Certificate
2001-01-12
2004-03-23
Marx, Irene (Department: 1651)
Chemistry: molecular biology and microbiology
Micro-organism, tissue cell culture or enzyme using process...
Preparing nitrogen-containing organic compound
C435S176000, C435S280000
Reexamination Certificate
active
06709847
ABSTRACT:
FIELD OF THE INVENTION
The present invention relates to an immobilized enzyme in which (S)-hydroxynitrile lyase is immobilized in an immobilization carrier with a high absorption ratio (or adsortion ratio), a method for producing said immobilized enzyme, and a method for producing optically active cyanohydrin using said immobilized enzyme.
BACKGROUND OF THE INVENTION
(S)-hydroxynitrile lyase is useful as an enzyme for synthesizing optically active cyanohydrins. In organic solvent reaction systems which are ordinarily used enzymatic synthesizing of the compounds, the enzyme are used, for example, as immobilized enzymes for dispersing the enzyme in the reaction system and for performing the reaction effectively. As an example in which (S)-hydroxynitrile lyase was immobilized, the immobilization to a micro cellulose powder and nitrocellulose has been reported. However, these cellulosic carriers have a low absorption ratio of the enzyme, arising inconveniences such that enormous amounts of carriers are necessary to immobilize the enzyme required for the reaction.
The object of the present invention is to provide an immobilized enzyme in which (S)-hydroxynitrile lyase is immobilized in an immobilization carrier at a high absorption ratio, a method for producing said immobilized enzyme, and a method for producing optically active cyanohydrin using said immobilized enzyme.
The inventors of the present invention have studied extensively and intensively to solve above problems and have now found that (S)-hydroxynitrile lyase could be immobilized at high absorption ratio by use of a porous inorganic carrier such as a sintered clay carrier, a silica carrier, an alumina carrier, and a silica alumina carrier as an immobilization carrier for enzyme, thereby completing the present invention.
The present invention relates to an immobilized enzyme in which (S)-hydroxynitrile lyase is immobilized in a carrier comprising a porous inorganic material (e.g. the sintered clay carrier, the silica carrier, the alumina carrier, or the silica alumina carrier, having a pore size of 10-80 nm). (S)-hydroxynitrile lyase used herein can be derived from a plant of Euphorbiaceae, Poaceae(Gramineae) or Olacaceae.
The present invention also relates to a method for producing the immobilized enzyme in which (S)-hydroxynitrile lyase is immobilized in a carrier comprising a porous inorganic material (e.g. the sintered clay carrier, the silica carrier, the alumina carrier or the silica alumina carrier, having the pore size of 10-80 nm). (S)-hydroxynitrile lyase used herein can be derived from a plant of Euphorbiaceae, Poaceae(Gramineae) or Olacaceae.
The present invention further relates to a method for producing an optically active cyanohydrin, comprising bringing said immobilized enzyme into contact with a carbonyl compound and a cyanogen compound in the presence of a slightly water-soluble or water-insoluble organic solvent. The immobilized enzyme used herein may be collected to be reused from the reaction mixture.
The present invention will be described in detail as follows.
REFERENCES:
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patent: 5885809 (1999-03-01), Effenberger
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Wajant, H. & Pfizenmaier, K., “Identification of Potential Active-site Redidues in the Hydroxynitrile Lyase fromManihot esculentaby Site-directed Mutagenesis,”J. Biol. Chem., 271(42): 25830-34 (1996).
E. Wehtje, et al. “Formation of C-C bonds by mandelonitrile lyase in organic solvents”Biotechnology and Bioengineering36: 39-46 (1990).
E. Wehtje, et al. “Activity and operational stability of immobilized mandelonitrile lyase in methanol/water mixtures”Appl. Microbiol. Biotechnol.29: 419-425 (1988).
E. Wehtje, et al. “Improved activity retention of enzymes deposited on solid supports”Biotechnology and Bioengineering41: 171-178 (1993).
Dobashi Yukio
Semba Hisashi
Finnegan Henderson Farabow Garrett & Dunner
Marx Irene
Nippon Shokubai Co. , Ltd.
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