The Elastase-PK101 Structure: Mechanism of an Ultrasensitive Activity-based Probe Revealed
Open accessAbstract
Human neutrophil elastase (HNE) plays a central role in neutrophil host defense, but its broad specificity makes HNE a difficult target for both inhibitor and probe development. Recently, we identified the unnatural amino acid containing activity-based probe PK101, which exhibits astounding sensitivity and selectivity for HNE, yet completely lacks mechanistic explanation for its unique characteristics. Here, we present the crystal structure of the HNE-PK101 complex which not only reveals the basis for PK101 ultrasensitivity but also uncovers so far unrecognized HNE features. Strikingly, the Nle(O-Bzl) function in the P4 position of PK101 reveals and leverages an "exo-pocket" on HNE as a critical factor for selectivity. Furthermore, the PK101 P3 position harbors a methionine dioxide function, which mimics a post-translationally oxidized methionine residue and forms a critical hydrogen bond to the backbone amide of Gly219 of HNE. Gly219 resides in a Gly-Gly motif that is unique to HNE, yet compulsory for this interaction. Consequently, this feature enables HNE to accommodate substrates that have undergone methionine oxidation, which constitutes a hallmark post-translational modification of neutrophil signaling.
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Crystallography of the PK101-HNE complex revealed an essential hydrogen bond between a sulfonyl oxygen of the oxidised methionine residue and the backbone amide of a unique Gly-Gly motif in the HNE S3 pocket.
“Subsequent crystallography to characterise the interaction between the screen-selected activity-based probe PK101 and the HNE binding surface revealed an essential hydrogen bond between a sulfonyl oxygen of the oxidised methionine residue and the backbone amide of a unique Gly-Gly motif in the HNE S3 pocket [21].”
From Discrimination of Methionine Sulfoxide and Sulfone by Human Neutrophil Elastase (2021) · cites this paper as [21]
Our kinetic data and molecular modelling predictions are consistent with an important role for P3 methionine sulfone substrates in HNE proteolytic activity enhancement via a mechanism similar to that described for PK-101.
“Our kinetic data and molecular modelling predictions are consistent with an important role for P3 methionine sulfone substrates in HNE proteolytic activity enhancement via a mechanism similar to that described for PK-101 [21].”
From Discrimination of Methionine Sulfoxide and Sulfone by Human Neutrophil Elastase (2021) · cites this paper as [21]
The crystal structure of the methionine sulfonyl-containing inhibitor PK-101 identified a hydrogen bond between its oxidised methionine residue and the main chain amide at Gly219 in the S3 pocket of HNE.
“The crystal structure of the methionine sulfonyl-containing inhibitor PK-101 [21] identified a hydrogen bond between the oxidised methionine residue present in PK-101 and the main chain amide at Gly219 in the S3 pocket of HNE.”
From Discrimination of Methionine Sulfoxide and Sulfone by Human Neutrophil Elastase (2021) · cites this paper as [21]
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Cited by
Sorted by citations
Discrimination of Methionine Sulfoxide and Sulfone by Human Neutrophil Elastase
Leahy, D., Grant, C., Jackson, A., Duff, A., Tardiota, N., Van Haeften, J., Chen, X., Peake, J. M., Kruppa, M. D., Smith, E. T., et al. (2021) · Molecules · doi:10.3390/molecules26175344 · Cited by 2 · see citing works
“Subsequent crystallography to characterise the interaction between the screen-selected activity-based probe PK101 and the HNE binding surface revealed an essential hydrogen bond between a sulfonyl oxygen of the oxidised methionine residue and the backbone amide of a unique Gly-Gly motif in the HNE S3 pocket [21].”
BackgroundMethodResultInfluential
- Authors
- Lechtenberg, B. C., Kasperkiewicz, P., Robinson, H., Drag, M., & Riedl, S. J.
- Affiliations
- Sanford Burnham Prebys Medical Discovery Institute (US)
- Wrocław University of Science and Technology (PL)
- Brookhaven National Laboratory (US)
- Division of Bioorganic Chemistry, Faculty of Chemistry, Wroclaw University of Technology, 50-370 Wroclaw, Poland
- Corresponding author
- Riedl, Stefan J.
- Venue
- ACS Chemical Biology
- Publisher
- American Chemical Society (ACS)
- Published
- 2015-01-22
- Volume
- 10
- Issue
- 4
- Pages
- 945-951
- ISSN
- 1554-8929 (ISSN-L)
- 1554-8937
- OpenAlex
- W1997474534
- Cited by
- 28
- References
- 0
- Citations by year
- 2026: 1 · 2023: 1 · 2022: 2 · 2021: 4 · 2020: 3 · 2019: 3 · 2018: 8 · 2017: 3 · 2016: 2 · 2015: 1
- FWCI
- 1.54
- Type
- journal article
- Language
- en
- Open access
- green
- www.osti.gov
- Locations
- ACS Chemical Biology (publishedVersion)
- PubMed (publishedVersion)
- OSTI OAI (U.S. Department of Energy Office of Scientific and Technical Information) (submittedVersion)
- Funders
- Biological and Environmental Research
- Sanford-Burnham Medical Research Institute
- National Center for Research Resources (P41RR012408)
- National Institute of General Medical Sciences (P41GM103473)
- European Molecular Biology Organization
- Basic Energy Sciences
- Topics
- Life Sciences › Biochemistry, Genetics and Molecular Biology › Cancer Research › Protease and Inhibitor Mechanisms
- Physical Sciences › Materials Science › Materials Chemistry › Enzyme Structure and Function
- Life Sciences › Pharmacology, Toxicology and Pharmaceutics › Toxicology › Organoselenium and organotellurium chemistry
- Keywords
- human neutrophil elastase, activity-based probes, crystal structure, unnatural amino acids, substrate specificity, methionine oxidation, hydrogen bonding, post-translational modification
- MeSH terms
- Humans, Hydrogen Bonding, Methionine, Oxidation-Reduction, Protein Conformation, Molecular Probes, Crystallography, X-Ray, Leukocyte Elastase
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