Large-Scale Production of Major House Dust Mite Allergen Der f2 Mutant (C8/119S) in Escherichia coli(MEDICAL BIOTECHNOLOGY)
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概要
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Hyposensitization, in which causative antigens of allergic diseases are injected, is the sole means of a radical cure for allergic diseases. Since the therapeutic allergens currently used are naturally extracted, producing preparations with a stable titer from such extracts is extremely difficult. There are several reports on the expression of recombinant mite allergens in Escherichia coli using inducers. The use of an inducer for industrial production will lead to high costs and, for therapeutic use, it must be removed in the purification process. C8/119S is a mutant of Der f 2, a major house dust mite allergen. The C8/119S gene was integrated downstream of the trp promoter to produce the expression plasmid (pWU11-C8/119S). Then, this expression plasmid was used to transform E. coli strain HB101 (pWU11-C8/119S/HB101). A recombinant E. coli pWU11-C8/1195/HB101 did not express C8/119S in a low-temperature culture (32℃), but C8/119S was induced to a high level of expression in a high-temperature culture (37℃). pWU11-C8/119S/HB101 proliferated when expression was induced by high temperature and an approximately 3-fold greater proliferation was obtained compared with the use of an inducer in a large-scale culture. The C8/119S protein was expressed as inclusion bodies and obtained by refolding and chromatography purifications. The immunological properties of C8/119S were assessed by western blotting. Western blotting demonstrated that purified C8/119S reacted with a monoclonal anti-Der f 2 antibody (18G8). pWU11-C8/119S/HB1O1 can be used as an easy, low cost expression system on a large scale. It is also advantageous for industrial production in that the addition of an inducer is not required to achieve expression of the mite allergen.
- 社団法人日本生物工学会の論文
- 2008-10-25
著者
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Murakami Toshio
Applied Research Department, The Chemo-Sero-Therapeutic Research Institute (Kaketsuken)
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Kawatsu Kenjirou
Applied Research Department, The Chemo-Sero-Therapeutic Research Institute (Kaketsuken)
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MAEDA Terunobu
Civil Chemical Engineering Corporation
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SUGAWARA Keishin
Applied Research Department, The Chemo-Sero-Therapeutic Research Institute (KAKETSUKEN)
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Maeda T
Civil Chemical Engineering Corporation
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Maeda Toshihiro
Applied Research Department The Chemo-sero-therapeutic Research Institute (kaketsuken)
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Koyanagi Satoshi
Applied Research Department, The Chemo-Sero-Therapeutic Research Institute (Kaketsuken)
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Miyatsu Yoshinobu
Applied Research Department, The Chemo-Sero-Therapeutic Research Institute (Kaketsuken)
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Mizokami Hiroshi
Applied Research Department, The Chemo-Sero-Therapeutic Research Institute (Kaketsuken)
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Sugawara Keishin
Biologics Development & Production Department The Chemo-sero-therapeutic Research Institute (kak
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Sugawara Keishin
Applied Research Department The Chemo-sero-therapeutic Research Institute (kaketsuken)
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Koyanagi Satoshi
Biologics Development & Production Department The Chemo-sero-therapeutic Research Institute (kak
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Kawatsu Kenjirou
Applied Research Department The Chemo-sero-therapeutic Research Institute (kaketsuken)
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Mizokami Hiroshi
Biologics Development & Production Department The Chemo-sero-therapeutic Research Institute (kak
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Mizokami Hiroshi
Department Of Research And Development
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Miyatsu Yoshinobu
Biologics Development & Production Department The Chemo-sero-therapeutic Research Institute (kak
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Murakami Toshio
Applied Research Department The Chemo-sero-therapeutic Research Institute (kaketsuken)
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