AU2002215754A1 - Compositions, methods and kits for identifying protein-protein interaction disrupting agents - Google Patents
Compositions, methods and kits for identifying protein-protein interaction disrupting agentsInfo
- Publication number
- AU2002215754A1 AU2002215754A1 AU2002215754A AU1575402A AU2002215754A1 AU 2002215754 A1 AU2002215754 A1 AU 2002215754A1 AU 2002215754 A AU2002215754 A AU 2002215754A AU 1575402 A AU1575402 A AU 1575402A AU 2002215754 A1 AU2002215754 A1 AU 2002215754A1
- Authority
- AU
- Australia
- Prior art keywords
- protein
- interaction
- drug
- hybrid system
- reverse
- 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.)
- Abandoned
Links
- 230000004850 protein–protein interaction Effects 0.000 title abstract 5
- 238000000034 method Methods 0.000 title abstract 2
- 239000000203 mixture Substances 0.000 title abstract 2
- 229940079593 drug Drugs 0.000 abstract 5
- 239000003814 drug Substances 0.000 abstract 5
- 230000003993 interaction Effects 0.000 abstract 3
- 238000010396 two-hybrid screening Methods 0.000 abstract 3
- 102000001301 EGF receptor Human genes 0.000 abstract 2
- 108060006698 EGF receptor Proteins 0.000 abstract 2
- 239000004098 Tetracycline Substances 0.000 abstract 2
- 239000003795 chemical substances by application Substances 0.000 abstract 2
- 230000002452 interceptive effect Effects 0.000 abstract 2
- 108090000623 proteins and genes Proteins 0.000 abstract 2
- 102000004169 proteins and genes Human genes 0.000 abstract 2
- 229960002180 tetracycline Drugs 0.000 abstract 2
- 229930101283 tetracycline Natural products 0.000 abstract 2
- 235000019364 tetracycline Nutrition 0.000 abstract 2
- 150000003522 tetracyclines Chemical class 0.000 abstract 2
- 108700008625 Reporter Genes Proteins 0.000 abstract 1
- 102000009661 Repressor Proteins Human genes 0.000 abstract 1
- 108010034634 Repressor Proteins Proteins 0.000 abstract 1
- 102000004887 Transforming Growth Factor beta Human genes 0.000 abstract 1
- 108090001012 Transforming Growth Factor beta Proteins 0.000 abstract 1
- 210000004027 cell Anatomy 0.000 abstract 1
- 238000004113 cell culture Methods 0.000 abstract 1
- 210000000349 chromosome Anatomy 0.000 abstract 1
- 150000001875 compounds Chemical class 0.000 abstract 1
- 239000002532 enzyme inhibitor Substances 0.000 abstract 1
- 229940125532 enzyme inhibitor Drugs 0.000 abstract 1
- 108020001507 fusion proteins Proteins 0.000 abstract 1
- 102000037865 fusion proteins Human genes 0.000 abstract 1
- 238000013537 high throughput screening Methods 0.000 abstract 1
- 239000000411 inducer Substances 0.000 abstract 1
- 230000002401 inhibitory effect Effects 0.000 abstract 1
- 229940043355 kinase inhibitor Drugs 0.000 abstract 1
- 210000004962 mammalian cell Anatomy 0.000 abstract 1
- 239000003757 phosphotransferase inhibitor Substances 0.000 abstract 1
- 239000013612 plasmid Substances 0.000 abstract 1
- 108020003175 receptors Proteins 0.000 abstract 1
- 102000005962 receptors Human genes 0.000 abstract 1
- 238000012216 screening Methods 0.000 abstract 1
- ZRKFYGHZFMAOKI-QMGMOQQFSA-N tgfbeta Chemical compound C([C@H](NC(=O)[C@H](C(C)C)NC(=O)CNC(=O)[C@H](CCC(O)=O)NC(=O)[C@H](CCCNC(N)=N)NC(=O)[C@H](CC(N)=O)NC(=O)[C@H](CC(C)C)NC(=O)[C@H]([C@@H](C)O)NC(=O)[C@H](CCC(O)=O)NC(=O)[C@H]([C@@H](C)O)NC(=O)[C@H](CC(C)C)NC(=O)CNC(=O)[C@H](C)NC(=O)[C@H](CO)NC(=O)[C@H](CCC(N)=O)NC(=O)[C@@H](NC(=O)[C@H](C)NC(=O)[C@H](C)NC(=O)[C@@H](NC(=O)[C@H](CC(C)C)NC(=O)[C@@H](N)CCSC)C(C)C)[C@@H](C)CC)C(=O)N[C@@H]([C@@H](C)O)C(=O)N[C@@H](C(C)C)C(=O)N[C@@H](CC=1C=CC=CC=1)C(=O)N[C@@H](C)C(=O)N1[C@@H](CCC1)C(=O)N[C@@H]([C@@H](C)O)C(=O)N[C@@H](CC(N)=O)C(=O)N[C@@H](CCC(O)=O)C(=O)N[C@@H](C)C(=O)N[C@@H](CC=1C=CC=CC=1)C(=O)N[C@@H](CCCNC(N)=N)C(=O)N[C@@H](C)C(=O)N[C@@H](CC(C)C)C(=O)N1[C@@H](CCC1)C(=O)N1[C@@H](CCC1)C(=O)N[C@@H](CCCNC(N)=N)C(=O)N[C@@H](CCC(O)=O)C(=O)N[C@@H](CCCNC(N)=N)C(=O)N[C@@H](CO)C(=O)N[C@@H](CCCNC(N)=N)C(=O)N[C@@H](CC(C)C)C(=O)N[C@@H](CC(C)C)C(O)=O)C1=CC=C(O)C=C1 ZRKFYGHZFMAOKI-QMGMOQQFSA-N 0.000 abstract 1
Classifications
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
- C12N15/00—Mutation or genetic engineering; DNA or RNA concerning genetic engineering, vectors, e.g. plasmids, or their isolation, preparation or purification; Use of hosts therefor
- C12N15/09—Recombinant DNA-technology
- C12N15/63—Introduction of foreign genetic material using vectors; Vectors; Use of hosts therefor; Regulation of expression
- C12N15/635—Externally inducible repressor mediated regulation of gene expression, e.g. tetR inducible by tetracyline
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
- C12N15/00—Mutation or genetic engineering; DNA or RNA concerning genetic engineering, vectors, e.g. plasmids, or their isolation, preparation or purification; Use of hosts therefor
- C12N15/09—Recombinant DNA-technology
- C12N15/10—Processes for the isolation, preparation or purification of DNA or RNA
- C12N15/1034—Isolating an individual clone by screening libraries
- C12N15/1055—Protein x Protein interaction, e.g. two hybrid selection
Landscapes
- Life Sciences & Earth Sciences (AREA)
- Health & Medical Sciences (AREA)
- Genetics & Genomics (AREA)
- Engineering & Computer Science (AREA)
- Chemical & Material Sciences (AREA)
- Organic Chemistry (AREA)
- Zoology (AREA)
- Wood Science & Technology (AREA)
- Biomedical Technology (AREA)
- Bioinformatics & Cheminformatics (AREA)
- Biotechnology (AREA)
- General Engineering & Computer Science (AREA)
- Molecular Biology (AREA)
- Plant Pathology (AREA)
- Biophysics (AREA)
- Microbiology (AREA)
- Physics & Mathematics (AREA)
- Biochemistry (AREA)
- General Health & Medical Sciences (AREA)
- Crystallography & Structural Chemistry (AREA)
- Bioinformatics & Computational Biology (AREA)
- Measuring Or Testing Involving Enzymes Or Micro-Organisms (AREA)
- Micro-Organisms Or Cultivation Processes Thereof (AREA)
- Investigating Or Analysing Biological Materials (AREA)
- Peptides Or Proteins (AREA)
Abstract
The present invention provides compositions, methods, and kits for identifying agents that are capable of disrupting protein-protein interaction in a mammalian reverse two-hybrid system. A tetracycline repressor protein was fused with the inhibitory KRAB domain as a suppressor to tightly regulate the reverse two-hybrid system for mammalian cells. Binding of the chimeric protein to the tetracycline operator sequence provided within a promoter entirely suppressed the expression of two interactive proteins. When both an inducer and a candidate protein-protein interaction disrupting agent such as a drug were added in the cell culture together, the reporter gene was either activated by the interaction of protein-pair if the drug was ineffective or remained silent due to the disruption of the protein-protein interaction by the effective drug. The plasmids for the suppressor and the reporter were integrated into chromosomes. Constructs for the expression of interactive proteins were either stably or transiently transfected into the cells. The utility of this system for screening drugs, particularly for enzyme inhibitor drugs, was demonstrated by using two well characterized interactions of the type I receptor for TGFbeta with FKPB12 and the EGF receptor with p85. The interaction between TGFbetaBI and FKPB12 or between EGF receptor and p85 were disrupted by FK506 and kinase inhibitor AG1478, respectively. The mammalian reverse two-hybrid system of the present invention can also be used for high throughput screening of compounds that disrupt protein-protein interactions.
Applications Claiming Priority (3)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| US25591000P | 2000-12-18 | 2000-12-18 | |
| US60255910 | 2000-12-18 | ||
| PCT/CA2001/001770 WO2002050259A2 (en) | 2000-12-18 | 2001-12-13 | Compositions, methods and kits for identifying protein-protein interaction disrupting agents |
Publications (1)
| Publication Number | Publication Date |
|---|---|
| AU2002215754A1 true AU2002215754A1 (en) | 2002-07-01 |
Family
ID=22970358
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| AU2002215754A Abandoned AU2002215754A1 (en) | 2000-12-18 | 2001-12-13 | Compositions, methods and kits for identifying protein-protein interaction disrupting agents |
Country Status (8)
| Country | Link |
|---|---|
| US (1) | US20040067483A1 (en) |
| EP (1) | EP1343882B1 (en) |
| AT (1) | ATE269899T1 (en) |
| AU (1) | AU2002215754A1 (en) |
| CA (1) | CA2431898A1 (en) |
| DE (1) | DE60104019T2 (en) |
| ES (1) | ES2223736T3 (en) |
| WO (1) | WO2002050259A2 (en) |
Families Citing this family (9)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| WO2002066504A2 (en) * | 2001-02-16 | 2002-08-29 | Hybrigenics | Protein-protein interactions in saccharomyces cerevisiae |
| EP1582590A1 (en) * | 2004-04-02 | 2005-10-05 | Aptanomics | Sequential intracellular screening method |
| JPWO2006062169A1 (en) * | 2004-12-08 | 2008-06-12 | 学校法人慶應義塾 | System for screening proteins / peptides localized in a specific region in a cell |
| US8282947B2 (en) | 2005-07-11 | 2012-10-09 | Make-up Art Cosmetics, Inc. | Low viscosity, unstable water-in-silicone emulsion cosmetic compositions and methods of use thereof |
| EP2044219B1 (en) * | 2006-06-30 | 2013-05-22 | DiscoveRx Corporation | Detectable nucleic acid tag |
| DE102008050702A1 (en) * | 2008-10-07 | 2010-04-15 | Thiesen, Hans-Juergen, Prof. | ChemBioNet component analysis using a stable cell-based TetR-KRAB luciferase detection system (TIS 10 cells) |
| CA2759211C (en) | 2009-04-24 | 2017-02-28 | Daniel Kelly Treiber | Assay for detecting protein by a cellular process |
| US20130035255A1 (en) * | 2010-03-26 | 2013-02-07 | Integratech Proteomics, Llc | Controlled release hybrid systems |
| GB201621891D0 (en) * | 2016-12-21 | 2017-02-01 | Autolus Ltd | Transcription system |
Family Cites Families (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| US5925523A (en) * | 1996-08-23 | 1999-07-20 | President & Fellows Of Harvard College | Intraction trap assay, reagents and uses thereof |
-
2001
- 2001-12-13 AU AU2002215754A patent/AU2002215754A1/en not_active Abandoned
- 2001-12-13 EP EP01271437A patent/EP1343882B1/en not_active Expired - Lifetime
- 2001-12-13 WO PCT/CA2001/001770 patent/WO2002050259A2/en not_active Application Discontinuation
- 2001-12-13 ES ES01271437T patent/ES2223736T3/en not_active Expired - Lifetime
- 2001-12-13 US US10/450,777 patent/US20040067483A1/en not_active Abandoned
- 2001-12-13 AT AT01271437T patent/ATE269899T1/en not_active IP Right Cessation
- 2001-12-13 DE DE60104019T patent/DE60104019T2/en not_active Expired - Fee Related
- 2001-12-13 CA CA002431898A patent/CA2431898A1/en not_active Abandoned
Also Published As
| Publication number | Publication date |
|---|---|
| WO2002050259A3 (en) | 2002-10-17 |
| EP1343882A2 (en) | 2003-09-17 |
| US20040067483A1 (en) | 2004-04-08 |
| CA2431898A1 (en) | 2002-06-27 |
| ES2223736T3 (en) | 2005-03-01 |
| DE60104019T2 (en) | 2005-07-14 |
| EP1343882B1 (en) | 2004-06-23 |
| ATE269899T1 (en) | 2004-07-15 |
| DE60104019D1 (en) | 2004-07-29 |
| WO2002050259A2 (en) | 2002-06-27 |
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