2022 Fiscal Year Final Research Report
Protein expression system inducible by low-toxic substances in Bacillus subtilis
Project/Area Number |
20K05797
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Research Category |
Grant-in-Aid for Scientific Research (C)
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Allocation Type | Multi-year Fund |
Section | 一般 |
Review Section |
Basic Section 38020:Applied microbiology-related
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Research Institution | Fukuyama University |
Principal Investigator |
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Project Period (FY) |
2020-04-01 – 2023-03-31
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Keywords | 枯草菌 / タンパク質発現 / フラボノイド / ラムノース / ペクチン / T7 RNAポリメラーゼ |
Outline of Final Research Achievements |
Protein expression systems inducible by low-toxicity substances were developed in Bacillus subtilis. The fusion of a flavonoid-responsive promoter (PqdoI) with the T7 RNA polymerase gene (T7 pol) was integrated into the chromosome, and a plasmid carrying the target gene under the control of the T7 promoter was introduced to create a flavonoid-inducible T7 expression system. Improving the PqdoI responsible for T7 pol control enhanced the expression level upon induction. T7 expression systems inducible sugars, such as rhamnose and pectin, were attempted to create by replacing the T7 pol control region with promoters responsive to those sugars. In the pectin-inducible system, induction required a long time, while in the rhamnose-inducible system, considerable expression leakage was observed even under the non-inducing condition.
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Free Research Field |
微生物学、分子生物学
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Academic Significance and Societal Importance of the Research Achievements |
枯草菌は哺乳類に対して非病原性であり、本研究で用いたいずれの誘導物質も哺乳類への毒性が低いので、当該T7発現系によって目的タンパク質の有害物質フリーでの獲得が容易となる。フラボノイド誘導型発現系では、厳密な発現制御と高発現量を両立できており、目的タンパク質が宿主細胞の増殖に悪影響を及ぼす場合に有用となる。アグリコン形態のフラボノイドは疎水性が高いので、特定の輸送体を介さずに膜透過で細胞内に取り込み可能である。したがって、当該発現系の他の細菌宿主への移行が比較的容易であると考えられる。誘導物質の1つであるケルセチンが安価なこともこの発現系の産業利用の可能性を高めている。
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