DK1027608T3 - Protein fragment complementation assays - Google Patents
Protein fragment complementation assaysInfo
- Publication number
- DK1027608T3 DK1027608T3 DK99936199.1T DK99936199T DK1027608T3 DK 1027608 T3 DK1027608 T3 DK 1027608T3 DK 99936199 T DK99936199 T DK 99936199T DK 1027608 T3 DK1027608 T3 DK 1027608T3
- Authority
- DK
- Denmark
- Prior art keywords
- protein
- dhfr
- fragment
- fragments
- interactions
- Prior art date
Links
Classifications
-
- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N33/00—Investigating or analysing materials by specific methods not covered by groups G01N1/00 - G01N31/00
- G01N33/48—Biological material, e.g. blood, urine; Haemocytometers
- G01N33/50—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing
- G01N33/68—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing involving proteins, peptides or amino acids
-
- 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
-
- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N33/00—Investigating or analysing materials by specific methods not covered by groups G01N1/00 - G01N31/00
- G01N33/48—Biological material, e.g. blood, urine; Haemocytometers
- G01N33/50—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing
- G01N33/53—Immunoassay; Biospecific binding assay; Materials therefor
- G01N33/566—Immunoassay; Biospecific binding assay; Materials therefor using specific carrier or receptor proteins as ligand binding reagents where possible specific carrier or receptor proteins are classified with their target compounds
-
- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N33/00—Investigating or analysing materials by specific methods not covered by groups G01N1/00 - G01N31/00
- G01N33/48—Biological material, e.g. blood, urine; Haemocytometers
- G01N33/50—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing
- G01N33/58—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing involving labelled substances
- G01N33/581—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing involving labelled substances with enzyme label (including co-enzymes, co-factors, enzyme inhibitors or substrates)
-
- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N2500/00—Screening for compounds of potential therapeutic value
Landscapes
- Life Sciences & Earth Sciences (AREA)
- Health & Medical Sciences (AREA)
- Engineering & Computer Science (AREA)
- Chemical & Material Sciences (AREA)
- Biomedical Technology (AREA)
- Molecular Biology (AREA)
- Immunology (AREA)
- Hematology (AREA)
- Urology & Nephrology (AREA)
- Biotechnology (AREA)
- Biochemistry (AREA)
- Microbiology (AREA)
- Physics & Mathematics (AREA)
- General Health & Medical Sciences (AREA)
- Analytical Chemistry (AREA)
- Genetics & Genomics (AREA)
- Food Science & Technology (AREA)
- Cell Biology (AREA)
- Medicinal Chemistry (AREA)
- General Physics & Mathematics (AREA)
- Pathology (AREA)
- Bioinformatics & Cheminformatics (AREA)
- Organic Chemistry (AREA)
- Zoology (AREA)
- General Engineering & Computer Science (AREA)
- Wood Science & Technology (AREA)
- Proteomics, Peptides & Aminoacids (AREA)
- Crystallography & Structural Chemistry (AREA)
- Bioinformatics & Computational Biology (AREA)
- Plant Pathology (AREA)
- Biophysics (AREA)
- Measuring Or Testing Involving Enzymes Or Micro-Organisms (AREA)
- Peptides Or Proteins (AREA)
- Investigating Or Analysing Biological Materials (AREA)
- Transition And Organic Metals Composition Catalysts For Addition Polymerization (AREA)
Abstract
We describe a strategy for designing and implementing protein-fragment complementation assays (PCAs) to detect biomolecular interactions in vivo and in vitro. A Protein Complementation Assay/Universal Reporter System (PCA/URS) for detecting and screening for agonists and antagonists of a membrane receptor is also described. The design, implementation and broad applications of this strategy are illustrated with a large number of enzymes with particular detail provided for the example of murine dihydrofolate reductase (DHFR). Fusion peptides consisting of N- and Cterminal fragments of murine DHFR fused to GCN4 leucine zipper sequences were coexpressed in Escherichia coli grown in minimal medium, where the endogenous DHFR activity was inhibited with trimethoprim. Coexpression of the complementary fusion products restored colony formation. Survival only occurred when both DHFR fragments were present and contained leucine-zipper forming sequences, demonstrating that reconstitution of enzyme activity requires assistance of leucine zipper formation. DHFR fragment-interface point mutants of increasing severity (Ile to Val, Ala and Gly) resulted in a sequential increase in E. coli doubling times illustrating the successful DHFR fragment reassembly rather than non-specific interactions between fragments. This assay could be used to study equilibrium and kinetic aspects of molecular interactions including protein-protein, protein-DNA, protein-RNA, protein-carbohydrate and protein-small molecule interactions, for screening cDNA libraries for binding of a target protein with unknown proteins or libraries of small organic molecules for biological activity. The selection and design criteria applied here is developed for numerous examples of clonal selection, colorometric, fluorometric and other assays based on enzymes whose products can be measured. The development of such assay systems is shown to be simple, and provides for a diverse set of protein fragment complementation applications.
Applications Claiming Priority (3)
Application Number | Priority Date | Filing Date | Title |
---|---|---|---|
CA002244349A CA2244349A1 (en) | 1998-07-30 | 1998-07-30 | Protein fragment complementation assays for the detection of biological or drug interactions |
US09/124,850 US6294330B1 (en) | 1997-01-31 | 1998-07-30 | Protein fragment complementation assays for the detection of biological or drug interactions |
PCT/CA1999/000702 WO2000007038A2 (en) | 1998-07-30 | 1999-07-30 | Protein fragment complementation assays |
Publications (1)
Publication Number | Publication Date |
---|---|
DK1027608T3 true DK1027608T3 (en) | 2010-05-25 |
Family
ID=29409708
Family Applications (1)
Application Number | Title | Priority Date | Filing Date |
---|---|---|---|
DK99936199.1T DK1027608T3 (en) | 1998-07-30 | 1999-07-30 | Protein fragment complementation assays |
Country Status (5)
Country | Link |
---|---|
AT (1) | ATE456054T1 (en) |
CA (1) | CA2244349A1 (en) |
DE (1) | DE69941946D1 (en) |
DK (1) | DK1027608T3 (en) |
ES (1) | ES2339411T3 (en) |
Families Citing this family (2)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
US7488583B2 (en) * | 2003-09-25 | 2009-02-10 | Odyssey Thera, Inc. | Fragment complementation assays for G-protein-coupled receptors and their signaling pathways |
CN114190538B (en) * | 2021-12-21 | 2023-11-24 | 郑州轻工业大学 | Grease crystallization promoter for promoting formation of beta' crystal form and application |
-
1998
- 1998-07-30 CA CA002244349A patent/CA2244349A1/en not_active Abandoned
-
1999
- 1999-07-30 AT AT99936199T patent/ATE456054T1/en not_active IP Right Cessation
- 1999-07-30 ES ES99936199T patent/ES2339411T3/en not_active Expired - Lifetime
- 1999-07-30 DK DK99936199.1T patent/DK1027608T3/en active
- 1999-07-30 DE DE69941946T patent/DE69941946D1/en not_active Expired - Lifetime
Also Published As
Publication number | Publication date |
---|---|
CA2244349A1 (en) | 2000-01-30 |
ATE456054T1 (en) | 2010-02-15 |
ES2339411T3 (en) | 2010-05-19 |
DE69941946D1 (en) | 2010-03-11 |
Similar Documents
Publication | Publication Date | Title |
---|---|---|
Titeca et al. | Discovering cellular protein‐protein interactions: Technological strategies and opportunities | |
DK0966685T3 (en) | Protein-fragment complementation assays to detect biomolecular interactions | |
AU2001278613B2 (en) | Functional protein arrays | |
Lamla et al. | The Nano-tag, a streptavidin-binding peptide for the purification and detection of recombinant proteins | |
Ohmuro-Matsuyama et al. | Homogeneous noncompetitive luminescent immunodetection of small molecules by ternary protein fragment complementation | |
US20030198971A1 (en) | Reactivation-based molecular interaction sensors | |
US10202466B2 (en) | Linked peptide fluorogenic biosensors | |
CA2384561A1 (en) | Methods of detecting interactions between proteins, peptides or libraries thereof using fusion proteins | |
Aggarwal et al. | Single‐molecule pull‐down (SiMPull) for new‐age biochemistry: Methodology and biochemical applications of single‐molecule pull‐down (SiMPull) for probing biomolecular interactions in crude cell extracts | |
WO2003062402A2 (en) | Use of collections of binding sites for sample profiling and other applications | |
US20160054331A1 (en) | Method for the detection and/or enrichment of analyte proteins and/or analyte peptides from a complex protein mixture | |
Stein et al. | Ultrasensitive scaffold-dependent protease sensors with large dynamic range | |
WO2000007038A3 (en) | Protein fragment complementation assays | |
Kast et al. | 3D structural information as a guide to protein engineering using genetic selection | |
Pelletier et al. | Mapping protein–protein interactions with combinatorial biology methods | |
Suwwan de Felipe et al. | Correlation between Ligand− Receptor Affinity and the Transcription Readout in a Yeast Three-Hybrid System | |
Pappas et al. | Direct interaction of the C-terminal domain of α-catenin and F-actin is necessary for stabilized cell-cell adhesion | |
CA2427747A1 (en) | Affinity maturation by competitive selection | |
US20060105407A1 (en) | Protein chip for analyzing interaction between protein and substrate peptide thereof | |
DK1027608T3 (en) | Protein fragment complementation assays | |
Ha et al. | Construction of allosteric protein switches by alternate frame folding and intermolecular fragment exchange | |
Groll et al. | Towards multiplexed protein–protein interaction analysis using protein tag-specific nanobodies | |
JP6624755B2 (en) | Protein tag, tagged protein and protein purification method | |
Kim et al. | On-chip Escherichia coli culture, purification, and detection of expressed proteins | |
JP6502854B2 (en) | Method of identifying polyubiquitinated substrate |