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Specificity profiling of human trypsin-isoenzymes

  • Oliver Schilling , Martin L. Biniossek , Bettina Mayer , Brigitta Elsässer , Hans Brandstetter , Peter Goettig ORCID logo , Ulf-Håkan Stenman und Hannu Koistinen EMAIL logo
Veröffentlicht/Copyright: 8. Juni 2018

Abstract

In humans, three different trypsin-isoenzymes have been described. Of these, trypsin-3 appears to be functionally different from the others. In order to systematically study the specificity of the trypsin-isoenzymes, we utilized proteome-derived peptide libraries and quantitative proteomics. We found similar specificity profiles dominated by the well-characterized preference for cleavage after lysine and arginine. Especially, trypsin-1 slightly favored lysine over arginine in this position, while trypsin-3 did not discriminate between them. In the P1′ position, which is the residue C-terminal to the cleavage site, we noticed a subtle enrichment of alanine and glycine for all three trypsins and for trypsin-3 there were additional minor P1′ and P2′ preferences for threonine and aspartic acid, respectively. These findings were confirmed by FRET peptide substrates showing different susceptibility to cleavage by different trypsins. The preference of trypsin-3 for aspartic acid in P2′ is explained by salt bridge formation with the unique Arg193. This salt bridge enables and stabilizes a canonical oxyanion conformation by the amides of Ser195 and Arg193, thus manifesting a selective substrate-assisted catalysis. As trypsin-3 has been proposed to be a therapeutic target and marker for cancers, our results may aid the development of specific inhibitors for cancer therapy and diagnostic probes.

Award Identifier / Grant number: SCHI 871/5, SCHI 871/8, SCHI 871/9, SCHI 871/11, INST 39/900-1, and SFB850-Project Z1

Award Identifier / Grant number: 1444

Funding statement: The authors thank Ms. Annikki Löfhjelm for excellent technical assistance. HK acknowledges support from the Finnish Cancer Foundation, Sigrid Jusélius Foundation and the Finnish Society of Clinical Chemistry. O.S. acknowledges support by Deutsche Forschungsgemeinschaft (SCHI 871/5, SCHI 871/8, SCHI 871/9, SCHI 871/11, INST 39/900-1, and SFB850-Project Z1), the Excellence Initiative of the German Federal and State Governments (EXC 294, BIOSS), the European Research Council (PoC 780730, ProteaseNter), and the German-Israeli Foundation for Scientific Research and Development (Funder Id: 10.13039/501100001736, grant no. 1444).

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Received: 2018-01-05
Accepted: 2018-04-05
Published Online: 2018-06-08
Published in Print: 2018-09-25

©2018 Walter de Gruyter GmbH, Berlin/Boston

Artikel in diesem Heft

  1. Frontmatter
  2. Highlight: The 7th International Symposium on Kallikreins and Kallikrein-Related Peptidases
  3. Obituary
  4. Manfred Schmitt (1947–2018)
  5. Functional interrelationships between the kallikrein-related peptidases family and the classical kinin system in the human neutrophil
  6. Overview of tissue kallikrein and kallikrein-related peptidases in breast cancer
  7. Kallikrein-related peptidases in lung diseases
  8. The miRNA-kallikrein interaction: a mosaic of epigenetic regulation in cancer
  9. Mining human cancer datasets for kallikrein expression in cancer: the ‘KLK-CANMAP’ Shiny web tool
  10. Specificity profiling of human trypsin-isoenzymes
  11. Activation and activity of glycosylated KLKs 3, 4 and 11
  12. Microenvironment proteinases, proteinase-activated receptor regulation, cancer and inflammation
  13. Kallikrein-related peptidase 6 orchestrates astrocyte form and function through proteinase activated receptor-dependent mechanisms
  14. Kallikrein-related peptidase 5 and seasonal influenza viruses, limitations of the experimental models for activating proteases
  15. Novel splice variants of the human kallikrein-related peptidases 11 (KLK11) and 12 (KLK12), unraveled by next-generation sequencing technology
  16. Insights into the activity control of the kallikrein-related peptidase 6: small-molecule modulators and allosterism
  17. Kallikrein-related peptidase 14 is the second KLK protease targeted by the serpin vaspin
  18. Profiling system for skin kallikrein proteolysis applied in gene-deficient mouse models
  19. Evidence that cell surface localization of serine protease activity facilitates cleavage of the protease activated receptor CDCP1
  20. Kallikrein-related peptidase 7 overexpression in melanoma cells modulates cell adhesion leading to a malignant phenotype
  21. KLK5, a novel potential suppressor of vaginal carcinogenesis
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