RU2017112965A - Слитые белки, рекомбинантные бактерии и способы применения рекомбинантных бактерий - Google Patents
Слитые белки, рекомбинантные бактерии и способы применения рекомбинантных бактерий Download PDFInfo
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- RU2017112965A RU2017112965A RU2017112965A RU2017112965A RU2017112965A RU 2017112965 A RU2017112965 A RU 2017112965A RU 2017112965 A RU2017112965 A RU 2017112965A RU 2017112965 A RU2017112965 A RU 2017112965A RU 2017112965 A RU2017112965 A RU 2017112965A
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- bacillus cereus
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- 241000894006 Bacteria Species 0.000 title 2
- 102000007056 Recombinant Fusion Proteins Human genes 0.000 title 1
- 108010008281 Recombinant Fusion Proteins Proteins 0.000 title 1
- 108090000623 proteins and genes Proteins 0.000 claims 61
- 241000193755 Bacillus cereus Species 0.000 claims 38
- 102000004169 proteins and genes Human genes 0.000 claims 34
- 230000035772 mutation Effects 0.000 claims 22
- 230000015572 biosynthetic process Effects 0.000 claims 6
- 102000034238 globular proteins Human genes 0.000 claims 5
- 108091005896 globular proteins Proteins 0.000 claims 5
- 108020001507 fusion proteins Proteins 0.000 claims 4
- 102000037865 fusion proteins Human genes 0.000 claims 4
- 210000004899 c-terminal region Anatomy 0.000 claims 3
- 241000654838 Exosporium Species 0.000 claims 2
- 108010043121 Green Fluorescent Proteins Proteins 0.000 claims 2
- 102000004144 Green Fluorescent Proteins Human genes 0.000 claims 2
- 239000012634 fragment Substances 0.000 claims 2
- 239000005090 green fluorescent protein Substances 0.000 claims 2
- 108090000765 processed proteins & peptides Proteins 0.000 claims 2
- 230000008685 targeting Effects 0.000 claims 2
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Claims (34)
1. Рекомбинантный представитель семейства Bacillus cereus, который экспрессирует слитый белок, содержащий по меньшей мере один представляющий интерес белок или пептид и нацеливающую последовательность, белок экзоспория или фрагмент белка экзоспория, которые нацеливают слитый белок в экзоспорий рекомбинантного представителя семейства Bacillus cereus; где рекомбинантный представитель семейства Bacillus cereus содержит мутацию или экспрессирует белок, где экспрессия белка увеличена по сравнению с экспрессией белка в представителе семейства Bacillus cereus дикого типа в тех же условиях, и где мутация или увеличенная экспрессия белка обеспечивает споры Bacillus cereus, имеющие экзоспорий, который легче удалить из споры по сравнению с экзоспорием споры дикого типа.
2. Рекомбинантный представитель семейства Bacillus cereus, который экспрессирует слитый белок, содержащий по меньшей мере один представляющий интерес белок или пептид и нацеливающую последовательность, белок экзоспория или фрагмент белка экзоспория, которые нацеливают слитый белок в экзоспорий рекомбинантного представителя семейства Bacillus cereus; где рекомбинантный представитель семейства Bacillus cereus:
(i) содержит мутацию в гене CotE;
(ii) эксрессирует белок ExsY, где экспрессия белка ExsY увеличена по сравнению с экспрессией белка ExsY в представителе семейства Bacillus cereus дикого типа в тех же условиях, и где белок ExsY содержит С-концевую метку, содержащую глобулярный белок;
(iii) экспрессирует белок BclB, где экспрессия белка BclB увеличена по сравнению с экспрессией белка BclB в представителе семейства Bacillus cereus дикого типа в тех же условиях;
(iv) экспрессирует белок YjcB, где экспрессия белка YjcB увеличена по сравнению с экспрессией белка YjcB в представителе семейства Bacillus cereus дикого типа в тех же условиях;
(v) содержит мутацию в гене ExsY;
(vi) содержит мутацию в гене CotY;
(vii) содержит мутацию в гене ExsA; или
(viii) содержит мутацию в гене CotO.
3. Рекомбинантный представитель семейства Bacillus cereus по п.1 или 2, где рекомбинантный представитель семейства Bacillus cereus содержит мутацию в гене CotE.
4. Рекомбинантный представитель семейства Bacillus cereus по п.3, где мутация в гене CotE включает нокаут гена CotE или доминантно-негативную форму гена CotE.
5. Рекомбинантный представитель семейства Bacillus cereus по любому из п.п.1-4, где рекомбинантный представитель семейства Bacillus cereus экспрессирует белок ExsY, где экспрессия белка ExsY увеличена по сравнению с экспрессией белка ExsY в представителе семейства Bacillus cereus дикого типа в тех же условиях, и где белок ExsY содержит С-концевую метку, содержащую глобулярный белок.
6. Рекомбинантный представитель семейства Bacillus cereus по п.5, где глобулярный белок имеет молекулярную массу от 25 кДа до 100 кДа.
7. Рекомбинантный представитель семейства Bacillus cereus по п.5 или 6, где глобулярный белок включает зеленый флуоресцентный белок (GFP) или его вариант.
8. Рекомбинантный представитель семейства Bacillus cereus по любому из п.п.1-7, где рекомбинантный представитель семейства Bacillus cereus экспрессирует белок BclB, где экспрессия белка BclB увеличена по сравнению с экспрессией белка BclB в представителе семейства Bacillus cereus дикого типа в тех же условиях.
9. Рекомбинантный представитель семейства Bacillus cereus по любому из п.п.1-8, где рекомбинантный представитель семейства Bacillus cereus экспрессирует белок YjcB, где экспрессия белка YjcB увеличена по сравнению с экспрессией белка YjcB в представителе семейства Bacillus cereus дикого типа в тех же условиях.
10. Рекомбинантный представитель семейства Bacillus cereus по любому из п.п.1-9, где рекомбинантный представитель семейства Bacillus cereus содержит мутацию в гене ExsY.
11. Рекомбинантный представитель семейства Bacillus cereus по п.10, где мутация в гене ExsY включает нокаут гена ExsY.
12. Рекомбинантный представитель семейства Bacillus cereus по любому из п.п.1-11, где рекомбинантный представитель семейства Bacillus cereus содержит мутацию в гене CotY.
13. Рекомбинантный представитель семейства Bacillus cereus по п.12, где мутация в гене CotY включает нокаут гена CotY.
14. Рекомбинантный представитель семейства Bacillus cereus по любому из п.п.1-13, где рекомбинантный представитель семейства Bacillus cereus содержит мутацию в гене ExsA.
15. Рекомбинантный представитель семейства Bacillus cereus по п.14, где мутация в гене ExsA включает нокаут гена ExsA.
16. Рекомбинантный представитель семейства Bacillus cereus по любому из п.п.1-15, где рекомбинантный представитель семейства Bacillus cereus содержит мутацию в гене CotO.
17. Рекомбинантный представитель семейства Bacillus cereus по п.16, где мутация в гене CotO включает нокаут гена CotO или доминантно-негативную форму гена CotO.
18. Рекомбинантный представитель семейства Bacillus cereus по любому из п.п.2-17, где:
мутация в гене CotE частично или полностью ингибирует способность CotE связывать экзоспорий со спорой;
экспрессия белка ExsY, содержащего С-концевую метку, содержащую глобулярный белок, ингибирует связывание белка ExsY с его мишенями в экзоспории;
экспрессия белка BclB приводит к образованию непрочного экзоспория;
экспрессия белка YjcB вызывает формирование экзоспория фрагментарно вместо полной структуры;
мутация в гене ExsY частично или полностью ингибирует способность ExsY к полному формированию экзоспория или связыванию экзоспория с спорой;
мутация в гене CotY приводит к образованию непрочного экзоспория;
мутация в гене ExsA приводит к формированию непрочного экзоспория; или
мутация в гене CotO вызывает формирование экзоспория полосами.
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US201462051885P | 2014-09-17 | 2014-09-17 | |
US62/051,885 | 2014-09-17 | ||
PCT/US2015/050807 WO2016044661A1 (en) | 2014-09-17 | 2015-09-17 | Fusion proteins, recombinant bacteria, and methods for using recombinant bacteria |
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RU2022111139A Division RU2022111139A (ru) | 2014-09-17 | 2015-09-17 | Слитые белки, рекомбинантные бактерии и способы применения рекомбинантных бактерий |
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