ArticleBiology open2014
Protein interference applications in cellular and developmental biology using DARPins that recognize GFP and mCherry.
Article in Biology open, 2014. The graph could read no effect estimate from its abstract, so it casts no vote on the map. Cited by 55 papers.
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Who cites it
55 citing papers in PubMed.
- Article
- Article
- CF2H: a cell-free two-hybrid platform for rapid protein binder screening.Nature communications · 2026Article
- Quantitative Degradation Rate Assessment of bioPROTACs Based on Peptide Degrons, E3 Domains, Adapters and Conjugated Small Molecules.ACS chemical biology · 2026Article
- A nucleotide-independent, pan-RAS-targeted DARPin elicits anti-tumor activity in a multimodal manner.Molecular oncology · 2025Article
- Design of Antigen-Targeting Fluorogenic Probes Utilizing Intramolecular Addition Reaction of Protein-Dye Hybrids.Journal of the American Chemical Society · 2025Article
- Molecular features defining the efficiency of bioPROTACs.Communications biology · 2025Article
- DARPin-induced reactivation of p53 in HPV-positive cells.Nature structural & molecular biology · 2025Article
- A modular cell-free protein biosensor platform using split T7 RNA polymerase.Science advances · 2025Article
- DARPins as a novel tool to detect and degrade p73.Cell death & disease · 2024Article
- Protein degradation kinetics measured by microinjection and live-cell fluorescence microscopy.Scientific reports · 2024Article
- A modular cell-free protein biosensor platform using split T7 RNA polymerase.bioRxiv : the preprint server for biology · 2024Article
- Lipid-mediated intracellular delivery of recombinant bioPROTACs for the rapid degradation of undruggable proteins.Nature communications · 2024Article
- Designed Ankyrin Repeat Proteins as Actin Labels of Distinct Cytoskeletal Structures in Living Cells.ACS nano · 2024Article
- Designed fluorescent protein cages as fiducial markers for targeted cell imaging.bioRxiv : the preprint server for biology · 2024Article
- A DARPin promotes faster onset of botulinum neurotoxin A1 action.Nature communications · 2023Article
- A bifunctional kinase-phosphatase module balances mitotic checkpoint strength and kinetochore-microtubule attachment stability.The EMBO journal · 2023Article
- Click display: a rapid and efficient in vitro protein display method for directed evolution.Nucleic acids research · 2023Article
- Nuclear myosin VI maintains replication fork stability.Nature communications · 2023Article
- Lipid Nanoparticle Delivery of Small Proteins for PotentACS applied materials & interfaces · 2023Article
Corrections and comments
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Authors and funding
9 authors.
Funding
No grant is acknowledged in the PubMed record.
Abstract
Protein-protein interactions are crucial for cellular homeostasis and play important roles in the dynamic execution of biological processes. While antibodies represent a well-established tool to study protein interactions of extracellular domains and secreted proteins, as well as in fixed and permeabilized cells, they usually cannot be functionally expressed in the cytoplasm of living cells. Non-immunoglobulin protein-binding scaffolds have been identified that also function intracellularly and are now being engineered for synthetic biology applications. Here we used the Designed Ankyrin Repeat Protein (DARPin) scaffold to generate binders to fluorescent proteins and used them to modify biological systems directly at the protein level. DARPins binding to GFP or mCherry were selected by ribosome display. For GFP, binders with KD as low as 160 pM were obtained, while for mCherry the best affinity was 6 nM. We then verified in cell culture their specific binding in a complex cellular environment and found an affinity cut-off in the mid-nanomolar region, above which binding is no longer detectable in the cell. Next, their binding properties were employed to change the localization of the respective fluorescent proteins within cells. Finally, we performed experiments in Drosophila melanogaster and Danio rerio and utilized these DARPins to either degrade or delocalize fluorescently tagged fusion proteins in developing organisms, and to phenocopy loss-of-function mutations. Specific protein binders can thus be selected in vitro and used to reprogram developmental systems in vivo directly at the protein level, thereby bypassing some limitations of approaches that function at the DNA or the RNA level.
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Read under generation 80e0d062 · epoch 390. Bibliography from PubMed, PubMed Central and OpenAlex; grants from NIH RePORTER; trial links from ClinicalTrials.gov; estimates, votes and beliefs from the OpenQuestion graph.