CA2410413A1 - Fluorescent proteins - Google Patents
Fluorescent proteins Download PDFInfo
- Publication number
- CA2410413A1 CA2410413A1 CA002410413A CA2410413A CA2410413A1 CA 2410413 A1 CA2410413 A1 CA 2410413A1 CA 002410413 A CA002410413 A CA 002410413A CA 2410413 A CA2410413 A CA 2410413A CA 2410413 A1 CA2410413 A1 CA 2410413A1
- Authority
- CA
- Canada
- Prior art keywords
- gfp
- fluorescent protein
- amino acid
- substituted
- protein according
- Prior art date
- Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
- Granted
Links
Classifications
-
- C—CHEMISTRY; METALLURGY
- C07—ORGANIC CHEMISTRY
- C07K—PEPTIDES
- C07K14/00—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof
- C07K14/435—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from animals; from humans
- C07K14/43504—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from animals; from humans from invertebrates
- C07K14/43595—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from animals; from humans from invertebrates from coelenteratae, e.g. medusae
-
- C—CHEMISTRY; METALLURGY
- C07—ORGANIC CHEMISTRY
- C07K—PEPTIDES
- C07K2319/00—Fusion polypeptide
Abstract
A GFP with an F64L mutation and E222G mutation is provided. This GFP has a bigger Stokes shift compared to other GFPs making it very suitable for high throughput screening due to a better resolution. This GFP also has an excitation maximum between the yellow GFP and the cyan GFP allowing for cleaner band separation when used together with those GFPs. Examples include the sequences in SEQ ID NOs: 3 and 4.
Claims (19)
1. A fluorescent protein derived from Green Fluorescent Protein (GFP) or any functional GFP analogue, wherein the amino acid in position 1 preceding the chromophore has been mutated and wherein the Glutamic acid in position 222 has been mutated said mutated GFP has an excitation maximum at a higher wavelength and the fluorescence is in-creased when the mutated GFP is expressed in cells incubated at a temperature of 30°C
or above compared to wild-type GFP.
or above compared to wild-type GFP.
2. A fluorescent protein according to the preceding claim, wherein the chromophore is in position 65-57 of the predicted primary amino acid sequence of GFP.
3. A fluorescent protein according to any one of the preceding claims, said protein being derived from Aequoria victorea or Renilla.
4. A fluorescent protein according to any one of the preceding claims, wherein the amino acid F in position 64 of the GFP has been substituted by an aliphatic amino acid.
5. A fluorescent protein according to any one of the preceding claims, wherein the amino acid F in position 64 of the GFP has been substituted by an amino acid selected from the group consisting of L, I, V, A and G.
6. A fluorescent protein according to any one of the preceding claims, wherein the amino acid F in position 64 of the GFP has been substituted by L.
7. A fluorescent protein according to any one of the preceding claims, wherein the amino acid E in position 222 of the GFP has been substituted by an amino acid selected from the group consisting of G, A, V, L, I, F, S, T, N, and Q.
8. A fluorescent protein according to any one of the preceding claims, wherein the amino acid E in position 222 of the GFP has been substituted by G.
9. A fluorescent protein according to any one of the preceding claims having the amino acid sequence disclosed in SEQ ID NO: 4.
10. A fluorescent protein according to any one of the preceding claims having the amino acid sequence disclosed in SEQ ID NO: 8.
11. A fusion compound consisting of a fluorescent protein (GFP) according to any of the preceding claims, wherein the GFP is linked to a polypeptide.
12. A fusion compound according to the preceding claim, wherein the polypeptide is a ki-nase, preferably the catalytic subunit of protein kinase A, or protein kinase C, or Erk1, or a cytoskeletal element.
13. A nucleotide sequence coding for the Fluorescent protein of any of the preceding claims.
14. A nucleotide sequence according to the preceding claim, shown in SEQ ID
NO: 3.
NO: 3.
15. A nucleotide sequence according to the preceding claim, shown in SEQ ID
NO: 7.
NO: 7.
16. A nucleotide sequence according to any of the preceding claims in the form of a DNA
sequence.
sequence.
17. A host transformed with a DNA construct according to any of the preceding claims.
18. A process for preparing a polypeptide, comprising cultivating a host according to any of the preceding claims and obtaining therefrom the polypeptide expressed by said nu-cleotide sequence.
19. Use of the fluorescent protein according to any of the preceding claims in an in vitro assay for measuring protein kinase activity, or dephosphorylation activity, or for measur-ing protein redistribution.
Applications Claiming Priority (9)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| DKPA200000953 | 2000-06-19 | ||
| DKPA200000953 | 2000-06-19 | ||
| US21268100P | 2000-06-20 | 2000-06-20 | |
| US60/212,681 | 2000-06-20 | ||
| US29017001P | 2001-05-10 | 2001-05-10 | |
| DKPA200100739 | 2001-05-10 | ||
| US60/290,170 | 2001-05-10 | ||
| DKPA200100739 | 2001-05-10 | ||
| PCT/EP2001/006848 WO2001098338A2 (en) | 2000-06-19 | 2001-06-18 | Fluorescent proteins |
Publications (2)
| Publication Number | Publication Date |
|---|---|
| CA2410413A1 true CA2410413A1 (en) | 2001-12-27 |
| CA2410413C CA2410413C (en) | 2010-05-18 |
Family
ID=27439814
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CA2410413A Expired - Lifetime CA2410413C (en) | 2000-06-19 | 2001-06-18 | Fluorescent proteins |
Country Status (11)
| Country | Link |
|---|---|
| US (2) | US7001986B2 (en) |
| EP (1) | EP1299414B1 (en) |
| JP (2) | JP2005518180A (en) |
| AT (1) | ATE439373T1 (en) |
| AU (2) | AU2001279669B2 (en) |
| CA (1) | CA2410413C (en) |
| DE (1) | DE60139556D1 (en) |
| DK (1) | DK1299414T3 (en) |
| ES (1) | ES2331109T3 (en) |
| IL (1) | IL153113A0 (en) |
| WO (1) | WO2001098338A2 (en) |
Families Citing this family (14)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| WO1998021355A1 (en) | 1996-11-15 | 1998-05-22 | Life Technologies, Inc. | Mutants of green fluorescent protein |
| GB0109858D0 (en) * | 2001-04-23 | 2001-06-13 | Amersham Pharm Biotech Uk Ltd | Fluorscent proteins |
| JP3829252B2 (en) | 2001-06-08 | 2006-10-04 | 独立行政法人理化学研究所 | Fluorescent protein |
| US7271241B2 (en) * | 2002-04-24 | 2007-09-18 | Los Alamos National Security, Llc | Directed evolution methods for improving polypeptide folding and solubility and superfolder fluorescent proteins generated thereby |
| DE03779067T1 (en) * | 2002-11-12 | 2006-06-22 | Zakrytoe Aktsionernoe Obschestvo "Evrogen" | FLUORESCENCE PROTEINS AND CHROMOPROTEINS FROM NON-AEQUOREA HYDROZOA SPECIES AND METHOD FOR USE THEREOF |
| CA2585231C (en) | 2003-10-24 | 2014-09-09 | The Regents Of The University Of California | Self-assembling split-fluorescent protein systems |
| CA2606876C (en) * | 2005-05-03 | 2018-07-10 | Georgia State University Research Foundation, Inc. | Fluorescent proteins and uses thereof |
| EP1785434A1 (en) | 2005-11-11 | 2007-05-16 | Ludwig-Maximilians-Universität München | Targeting and tracing of antigens in living cells |
| EP2054514A4 (en) | 2006-08-04 | 2009-11-04 | Univ Georgia State Res Found | Enzyme sensors, methods for preparing and using such sensors, and methods of detecting protease activity |
| EP2189528A4 (en) * | 2007-08-03 | 2010-09-22 | Univ Hokkaido Nat Univ Corp | Fluorescent protein with deep blue color |
| EP2078750B1 (en) | 2008-01-09 | 2016-09-28 | Ludwig-Maximilians-Universität München | A fluorescent two-hybrid (F2H) assay for direct visualization of protein interactions in living cells |
| EP2275442A1 (en) | 2009-07-06 | 2011-01-19 | Ludwig-Maximilians-Universität München | Detection and vizualization of the cell cycle in living cells |
| CN101974554A (en) * | 2010-09-26 | 2011-02-16 | 武汉大学 | Construction method and application of mutant vector pEGFP-N1m |
| EP2685260A1 (en) | 2012-07-09 | 2014-01-15 | Ludwig-Maximilians-Universität München | Direct and quantitative detection of targets in living cells |
Family Cites Families (10)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| EP0759170B1 (en) | 1993-09-10 | 2008-07-09 | The Trustees Of Columbia University In The City Of New York | Uses of green fluorescent protein |
| US5491084A (en) * | 1993-09-10 | 1996-02-13 | The Trustees Of Columbia University In The City Of New York | Uses of green-fluorescent protein |
| US5610031A (en) * | 1993-10-27 | 1997-03-11 | The General Hospital Corporation | B1k chain of laminin and methods of use |
| WO1995021191A1 (en) | 1994-02-04 | 1995-08-10 | William Ward | Bioluminescent indicator based upon the expression of a gene for a modified green-fluorescent protein |
| US5625048A (en) * | 1994-11-10 | 1997-04-29 | The Regents Of The University Of California | Modified green fluorescent proteins |
| DE69604298T2 (en) * | 1995-09-22 | 2000-05-18 | Bioimage A S Soeborg | VARIANTS OF THE GREEN FLUORESCENCE PROTEIN, GFP |
| US5804387A (en) * | 1996-02-01 | 1998-09-08 | The Board Of Trustees Of The Leland Stanford Junior University | FACS-optimized mutants of the green fluorescent protein (GFP) |
| US6124128A (en) * | 1996-08-16 | 2000-09-26 | The Regents Of The University Of California | Long wavelength engineered fluorescent proteins |
| US6090919A (en) * | 1997-01-31 | 2000-07-18 | The Board Of Trustees Of The Leland Stanford Junior University | FACS-optimized mutants of the green fluorescent protein (GFP) |
| GB0109858D0 (en) | 2001-04-23 | 2001-06-13 | Amersham Pharm Biotech Uk Ltd | Fluorscent proteins |
-
2001
- 2001-06-18 EP EP01957861A patent/EP1299414B1/en not_active Expired - Lifetime
- 2001-06-18 IL IL15311301A patent/IL153113A0/en unknown
- 2001-06-18 CA CA2410413A patent/CA2410413C/en not_active Expired - Lifetime
- 2001-06-18 AU AU2001279669A patent/AU2001279669B2/en not_active Expired
- 2001-06-18 DK DK01957861T patent/DK1299414T3/en active
- 2001-06-18 DE DE60139556T patent/DE60139556D1/en not_active Expired - Lifetime
- 2001-06-18 ES ES01957861T patent/ES2331109T3/en not_active Expired - Lifetime
- 2001-06-18 WO PCT/EP2001/006848 patent/WO2001098338A2/en active IP Right Grant
- 2001-06-18 JP JP2002504293A patent/JP2005518180A/en not_active Withdrawn
- 2001-06-18 AT AT01957861T patent/ATE439373T1/en not_active IP Right Cessation
- 2001-06-18 AU AU7966901A patent/AU7966901A/en active Pending
- 2001-06-19 US US09/887,784 patent/US7001986B2/en not_active Expired - Lifetime
-
2005
- 2005-08-19 US US11/206,904 patent/US7476518B2/en not_active Expired - Lifetime
-
2006
- 2006-11-09 JP JP2006304095A patent/JP4459944B2/en not_active Expired - Lifetime
Also Published As
| Publication number | Publication date |
|---|---|
| WO2001098338A2 (en) | 2001-12-27 |
| WO2001098338A3 (en) | 2002-05-10 |
| DE60139556D1 (en) | 2009-09-24 |
| US20060051843A1 (en) | 2006-03-09 |
| ATE439373T1 (en) | 2009-08-15 |
| JP4459944B2 (en) | 2010-04-28 |
| AU2001279669B2 (en) | 2006-02-02 |
| US20020177189A1 (en) | 2002-11-28 |
| US7476518B2 (en) | 2009-01-13 |
| DK1299414T3 (en) | 2009-11-30 |
| AU7966901A (en) | 2002-01-02 |
| EP1299414B1 (en) | 2009-08-12 |
| JP2007075122A (en) | 2007-03-29 |
| US7001986B2 (en) | 2006-02-21 |
| ES2331109T3 (en) | 2009-12-22 |
| IL153113A0 (en) | 2003-06-24 |
| JP2005518180A (en) | 2005-06-23 |
| EP1299414A2 (en) | 2003-04-09 |
| CA2410413C (en) | 2010-05-18 |
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Legal Events
| Date | Code | Title | Description |
|---|---|---|---|
| EEER | Examination request | ||
| MKEX | Expiry |
Effective date: 20210618 |