ArticlePloS one2019
Enhanced trypsin on a budget: Stabilization, purification and high-temperature application of inexpensive commercial trypsin for proteomics applications.
Article in PloS one, 2019. The graph could read no effect estimate from its abstract, so it casts no vote on the map. Cited by 17 papers.
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Who cites it
17 citing papers in PubMed.
- Single-cell and spatial transcriptomics reveals the spatiotemporal trajectory of the small peptide TAP4 in delaying postharvest fruit senescence.Plant communications · 2026Article
- A posttranslational proteomic survey of a single anatomically preserved human 20-week postconception brain.Journal of anatomy · 2026Article
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- Challenges and Insights in Absolute Quantification of Recombinant Therapeutic Antibodies by Mass Spectrometry: An Introductory Review.Antibodies (Basel, Switzerland) · 2025Review
- Clean and Complete Protein Digestion with an Autolysis Resistant Trypsin for Peptide Mapping.Journal of proteome research · 2024Article
- A Mass Spectrometry Strategy for Protein Quantification Based on the Differential Alkylation of Cysteines Using Iodoacetamide and Acrylamide.International journal of molecular sciences · 2024Article
- Article
- Review
- Protein Digestion for 2D-DIGE Analysis.Methods in molecular biology (Clifton, N.J.) · 2023Article
- Detergent-Assisted Protein Digestion-On the Way to Avoid the Key Bottleneck of Shotgun Bottom-Up Proteomics.International journal of molecular sciences · 2022Review
- Review
- Maximizing Cumulative Trypsin Activity with Calcium at Elevated Temperature for Enhanced Bottom-Up Proteome Analysis.Biology · 2022Article
- Bottom-Up Analysis of Proteins by Peptide Mass Fingerprinting with tCITP-CZE-ESI-TOF MS After Tryptic Digest.Methods in molecular biology (Clifton, N.J.) · 2022Article
- Conjugation with gold nanoparticles improves the stability of the KT2 peptide and maintains its anticancer properties.RSC advances · 2021Article
- Article
- Citrullinome ofMolecular & cellular proteomics : MCP · 2020Article
- Nonspecific cleavages arising from reconstitution of trypsin under mildly acidic conditions.PloS one · 2020Article
Corrections and comments
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Authors and funding
4 authors.
Funding
No grant is acknowledged in the PubMed record.
Abstract
Trypsin is by far the most commonly used protease in proteomics. Even though the amount of protease used in each experiment is very small, digestion of large amounts of protein prior to enrichment can be rather costly. The price of commercial trypsin is highly dependent on the quality of the enzyme, which is determined by its purity, activity, and chemical modifications. In this study we evaluated several strategies for improving the quality of crude trypsin by reductive methylation and affinity purification. We present a protocol applicable to most proteomics laboratories for obtaining a highly stable and pure trypsin preparation using reductive methylation and purification by benzamidine-sepharose. The entire workflow can be performed within a day and yields ~4 mg per batch but is completely scalable. The methylated product was benchmarked against sequencing grade trypsin from Promega and they were found to be comparable for one hour digestions at elevated temperatures, where residual chymotryptic activity was found to be negligible.
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