ArticleACS chemical biology2016
Intracellular FRET-based Screen for Redesigning the Specificity of Secreted Proteases.
Article in ACS chemical biology, 2016. The graph could read no effect estimate from its abstract, so it casts no vote on the map. Cited by 20 papers.
What it found
Each row is one number read from the abstract, on the scale the paper reported it, with its interval. Left of the dashed line favours the treatment, right favours the comparator. Under each row is the sentence it came from. New to these charts? A ten-minute tutorial.
The abstract states no effect estimate the extractor could read, or names no intervention and outcome on the map, so this paper lights no cell and moves no belief. It is still indexed, cited and linked below.
The trial behind it
Trials whose registry record cites this paper, or whose number appears in the abstract. A trial that started after this paper was published is citing it as background, not reporting it.
Neither the registry nor the abstract names a trial number. If this is a trial report, that itself is worth knowing.
Who cites it
20 citing papers in PubMed, 33 citations in OpenAlex.
- Protease engineering: Approaches, tools, and emerging trends.Biotechnology advances · 2025Review
- Machine-guided dual-objective protein engineering for deimmunization and therapeutic functions.Cell systems · 2025Article
- Leveraging yeast sequestration to study and engineer posttranslational modification enzymes.Biotechnology and bioengineering · 2024Review
- Modular and integrative activity reporters enhance biochemical studies in the yeast ER.Protein engineering, design & selection : PEDS · 2024Article
- A high-throughput screening platform for enzymes active on mucin-type O-glycoproteins.Nature chemical biology · 2023Article
- Evolution of protease activation and specificity via alpha-2-macroglobulin-mediated covalent capture.Nature communications · 2023Article
- Reprogramming the Cleavage Specificity of Botulinum Neurotoxin Serotype B1.ACS synthetic biology · 2022Article
- Reengineering the specificity of the highly selective Clostridium botulinum protease via directed evolution.Scientific reports · 2022Article
- Making the cut with protease engineering.Cell chemical biology · 2022Review
- The past, present, and future of enzyme-based therapies.Drug discovery today · 2022Review
- Phage-assisted evolution of botulinum neurotoxin proteases with reprogrammed specificity.Science (New York, N.Y.) · 2021Article
- FRET Microscopy in Yeast.Biosensors · 2019Review
- Article
- Multiple domains of bacterial and human Lon proteases define substrate selectivity.Emerging microbes & infections · 2018Article
- Aminopeptidase secreted by Chromobacterium sp. Panama inhibits dengue virus infection by degrading the E protein.PLoS neglected tropical diseases · 2018Article
- Specificity of peptidases secreted by filamentous fungi.Bioengineered · 2018Review
- Characterization of aromatic residue-controlled protein retention in the endoplasmic reticulum ofThe Journal of biological chemistry · 2017Article
- Phage-assisted continuous evolution of proteases with altered substrate specificity.Nature communications · 2017Article
- Mass spectrometry-based determination of Kallikrein-related peptidase 7 (KLK7) cleavage preferences and subsite dependency.Scientific reports · 2017Article
- Evolution of a mass spectrometry-grade protease with PTM-directed specificity.Proceedings of the National Academy of Sciences of the United States of America · 2016Article
Corrections and comments
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Authors and funding
3 authors at 1 institution in 1 country.
Funding
Abstract
Proteases are attractive as therapeutics given their ability to catalytically activate or inactivate their targets. However, therapeutic use of proteases is limited by insufficient substrate specificity, since off-target activity can induce undesired side-effects. In addition, few methods exist to enhance the activity and specificity of human proteases, analogous to methods for antibody engineering. Given this need, a general methodology termed protease evolution via cleavage of an intracellular substrate (PrECISE) was developed to enable engineering of human protease activity and specificity toward an arbitrary peptide target. PrECISE relies on coexpression of a protease and a peptide substrate exhibiting Förster resonance energy transfer (FRET) within the endoplasmic reticulum of yeast. Use of the FRET reporter substrate enabled screening large protease libraries using fluorescence activated cell sorting for the activity of interest. To evolve a human protease that selectively cleaves within the central hydrophobic core (KLVF↓F↓AED) of the amyloid beta (Aβ) peptide, PrECISE was applied to human kallikrein 7, a protease with Aβ cleavage activity but broad selectivity, with a strong preference for tyrosine (Y) at P1. This method yielded a protease variant which displayed up to 30-fold improvements in Aβ selectivity mediated by a reduction in activity toward substrates containing tyrosine. Additionally, the increased selectivity of the variant led to reduced toxicity toward PC12 neuronal-like cells and 16-1000-fold improved resistance to wild-type inhibitors. PrECISE thus provides a powerful high-throughput capability to redesign human proteases for therapeutic use.
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Registered trials
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.