RU2208639C2 - Секреторная сигнальная последовательность stii (варианты), кодирующая ее нуклеиновая кислота, плазмидный вектор, способ оптимизации секреции и способ секреции гетерологического полипептида - Google Patents
Секреторная сигнальная последовательность stii (варианты), кодирующая ее нуклеиновая кислота, плазмидный вектор, способ оптимизации секреции и способ секреции гетерологического полипептида Download PDFInfo
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Abstract
Изобретение относится к генетической инженерии. Представлены варианты нуклеиновых кислот, содержащих секреторную сигнальную последовательность ST II. Нуклеотидные последовательности приведены в описании. Сравнивают уровни экспрессии полипептида, регулируемого набором вариантов нуклеиновых кислот, кодирующие секреторную сигнальную последовательность ST II. Выявляют варианты, имеющие эффективность трансляции, которая определяет оптимальную секрецию зрелого полипептида. Прокариотическую клетку хозяина трансформируют вектором, включающим секреторную сигнальную последовательность ST II, функционально присоединенную к нуклеиновой кислоте, кодирующей целевой гетерологический полипептид. Изобретение позволяет обеспечить при снижении трансляционной эффективности полное превращение предшественника в зрелый продукт и высокие уровни секреции целевого полипептида. 13 с. и 19 з.п.ф-лы, 2 табл., 21 ил.
Description
Текст описания в факсимильном виде (см. графическую часть). Т
Claims (32)
1. Нуклеиновая кислота, представляющая собой вариант нуклеиновой кислоты, кодирующий секреторную сигнальную последовательность STII, имеющий нуклеотидную последовательность, выбранную из группы, состоящей из SEQ ID :15 - SEQ ID :23.
2. Нуклеиновая кислота по п.1, отличающаяся тем, что указанный вариант не изменяет аминокислотную последовательность указанной секреторной сигнальной последовательности STII.
3. Нуклеиновая кислота по п.1, отличающаяся тем, что количество секретируемого гетерологического полипептида в случае, когда нуклеиновая кислота, кодирующая указанный гетерологический полипептид, функционально присоединена к указанному варианту и экспрессируется в клетке-хозяине больше, чем количество секретируемого гетерологического полипептида в случае, когда указанная нуклеиновая кислота, кодирующая указанный гетерологический полипептид, функционально присоединена к нуклеиновой кислоте дикого типа, кодирующей указанную секреторную сигнальную последовательность STII.
4. Нуклеиновая кислота по п.1, отличающаяся тем, что указанная секреторная сигнальная последовательность является прокариотической секреторной сигнальной последовательностью.
5. Нуклеиновая кислота по п.1, отличающаяся тем, характеризуется нуклеотидной последовательностью SEQ ID :15.
6. Нуклеиновая кислота по п.1, отличающаяся тем, что характеризуется нуклеотидной последовательностью SEQ ID :16.
7. Нуклеиновая кислота по п.1, отличающаяся тем, что характеризуется нуклеотидной последовательностью SEQ ID :17.
8. Нуклеиновая кислота по п.1, отличающаяся тем, что характеризуется нуклеотидной последовательностью SEQ ID :18.
9. Нуклеиновая кислота по п.1, отличающаяся тем, что характеризуется нуклеотидной последовательностью SEQ ID :19.
10. Нуклеиновая кислота по п.1, отличающаяся тем, что характеризуется нуклеотидной последовательностью SEQ ID :20.
11. Нуклеиновая кислота по п.1, отличающаяся тем, что характеризуется нуклеотидной последовательностью SEQ ID :21.
12. Нуклеиновая кислота по п.1, отличающаяся тем, что характеризуется нуклеотидной последовательностью SEQ ID :22.
13. Нуклеиновая кислота по п.1, отличающаяся тем, что характеризуется нуклеотидной последовательностью SEQ ID :23.
14. Способ оптимизации секреции интересующего гетерологического полипептида в прокариотической клетке, включающий сравнение уровней экспрессии полипептида, регулируемого набором вариантов нуклеиновой кислоты по п.1, при том, что набор вариантов представляет серию эффективностей трансляции, и идентификацию одного или более вариантов, имеющих эффективность трансляции, которая определяет оптимальную секрецию зрелого полипептида.
15. Способ по п.14, отличающийся тем, что указанный один или более идентифицированный вариант не изменяет аминокислотную последовательность указанной секреторной сигнальной последовательности STII.
16. Способ по п.14, отличающийся тем, что количество секретируемого полипептида в случае, когда нуклеиновая кислота, кодирующая указанный полипептид, функционально присоединена по крайней мере к одному из указанных идентифицированных вариантов, больше, чем количество секретируемого полипептида в случае, когда указанная нуклеиновая кислота функционально присоединена к нуклеиновой кислоте дикого типа, кодирующей указанную секреторную сигнальную последовательность STII.
17. Способ по п.14, отличающийся тем, что указанная секреторная сигнальная последовательность является прокариотической секреторной сигнальной последовательностью.
18. Способ секреции интересующего гетерологического полипептида в прокариотической клетке, включающий трансформацию прокариотической клетки-хозяина вектором, включающим нуклеиновую кислоту по п.1, функционально присоединенную к нуклеиновой кислоте, кодирующей указанный гетерологический полипептид, с целью секреции указанного гетерологического полипептида, при том, что указанный гетерологический полипептид секретируется.
19. Способ по п.18, отличающийся тем, что указанный вариант не изменяет аминокислотную последовательность указанной секреторной сигнальной последовательности STII.
20. Способ по п.18, отличающийся тем, что количество секретируемого полипептида в случае, когда указанная нуклеиновая кислота, кодирующая указанный гетерологический полипептид, функционально присоединена к указанной нуклеиновой кислоте, кодирующей вариант секреторной сигнальной последовательности STII, больше, чем количество секретируемого полипептида в случае, когда указанная нуклеиновая кислота, кодирующая указанный гетерологический полипептид, функционально присоединена к нуклеиновой кислоте дикого типа, кодирующей указанную секреторную сигнальную последовательность STII.
21. Способ по п. 18, отличающийся тем, что указанная секреторная сигнальная последовательность является прокариотической секреторной сигнальной последовательностью.
22. Плазмидный вектор, включающий нуклеиновую кислоту по любому из пп. 1-13, функционально присоединенную к дополнительным элементам, для осуществления экспрессии интересующего гена.
23. Вектор по п.22, используемый для встраивания в клетку-хозяин.
24. Вариант секреторной сигнальной последовательности STII, кодируемый нуклеотидной последовательностью SEQ ID :15.
25. Вариант секреторной сигнальной последовательности STII, кодируемый нуклеотидной последовательностью SEQ ID :16.
26. Вариант секреторной сигнальной последовательности STII, кодируемый нуклеотидной последовательностью SEQ ID :17.
27. Вариант секреторной сигнальной последовательности STII, кодируемый нуклеотидной последовательностью SEQ ID :18.
28. Вариант секреторной сигнальной последовательности STII, кодируемый нуклеотидной последовательностью SEQ ID :19.
29. Вариант секреторной сигнальной последовательности STII, кодируемый нуклеотидной последовательностью SEQ ID :20.
30. Вариант секреторной сигнальной последовательности STII, кодируемый нуклеотидной последовательностью SEQ ID :21.
31. Вариант секреторной сигнальной последовательности, кодируемый нуклеотидной последовательностью SEQ ID :22.
32. Вариант секреторной сигнальной последовательности STII, кодируемый нуклеотидной последовательностью SEQ ID :23.
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US08/398,615 | 1995-03-01 | ||
US08/398,615 US5840523A (en) | 1995-03-01 | 1995-03-01 | Methods and compositions for secretion of heterologous polypeptides |
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RU2208639C2 true RU2208639C2 (ru) | 2003-07-20 |
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JP (1) | JP3375970B2 (ru) |
CN (1) | CN1154736C (ru) |
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AU (1) | AU701476B2 (ru) |
BR (1) | BR9607464B1 (ru) |
CA (1) | CA2213813C (ru) |
DE (1) | DE69636996T2 (ru) |
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FI (1) | FI120357B (ru) |
IL (1) | IL117314A0 (ru) |
NO (1) | NO324920B1 (ru) |
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BR9607464B1 (pt) | 2009-01-13 |
CN1180379A (zh) | 1998-04-29 |
IL117314A0 (en) | 1996-06-18 |
CA2213813C (en) | 2008-05-27 |
MX9706607A (es) | 1997-11-29 |
NO324920B1 (no) | 2008-01-07 |
DE69636996T2 (de) | 2007-12-06 |
EP0815246B1 (en) | 2007-03-28 |
NZ303619A (en) | 2000-02-28 |
AU701476B2 (en) | 1999-01-28 |
JPH10507368A (ja) | 1998-07-21 |
CN1154736C (zh) | 2004-06-23 |
ATE358181T1 (de) | 2007-04-15 |
DK0815246T3 (da) | 2007-08-06 |
NO973985L (no) | 1997-10-29 |
NO973985D0 (no) | 1997-08-29 |
JP3375970B2 (ja) | 2003-02-10 |
ES2285711T3 (es) | 2007-11-16 |
CA2213813A1 (en) | 1996-09-06 |
DE69636996D1 (de) | 2007-05-10 |
US5840523A (en) | 1998-11-24 |
FI973559A (fi) | 1997-10-29 |
WO1996027016A1 (en) | 1996-09-06 |
ZA961688B (en) | 1997-09-01 |
FI120357B (fi) | 2009-09-30 |
FI973559A0 (fi) | 1997-08-29 |
BR9607464A (pt) | 1997-12-23 |
AU4998496A (en) | 1996-09-18 |
EP0815246A1 (en) | 1998-01-07 |
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