Engineered protease inhibitors based on sunflower trypsin inhibitor-1 (SFTI-1) provide insights into the role of sequence and conformation in Laskowski mechanism inhibition

de Veer, Simon J., Swedberg, Joakim E., Akcan, Muharrem, Rosengren, K. Johan, Brattsand, Maria, Craik, David J. and Harris, Jonathan M. (2015) Engineered protease inhibitors based on sunflower trypsin inhibitor-1 (SFTI-1) provide insights into the role of sequence and conformation in Laskowski mechanism inhibition. Biochemical Journal, 469 2: 243-253. doi:10.1042/BJ20150412


Author de Veer, Simon J.
Swedberg, Joakim E.
Akcan, Muharrem
Rosengren, K. Johan
Brattsand, Maria
Craik, David J.
Harris, Jonathan M.
Title Engineered protease inhibitors based on sunflower trypsin inhibitor-1 (SFTI-1) provide insights into the role of sequence and conformation in Laskowski mechanism inhibition
Journal name Biochemical Journal   Check publisher's open access policy
ISSN 1470-8728
0264-6021
Publication date 2015-06-06
Sub-type Article (original research)
DOI 10.1042/BJ20150412
Open Access Status Not yet assessed
Volume 469
Issue 2
Start page 243
End page 253
Total pages 11
Place of publication London, United Kingdom
Publisher Portland Press
Collection year 2016
Language eng
Abstract Laskowski inhibitors regulate serine proteases by an intriguing mode of action that involves deceiving the protease into synthesizing a peptide bond. Studies exploring naturally occurring Laskowski inhibitors have uncovered several structural features that convey the inhibitor's resistance to hydrolysis and exceptional binding affinity. However, in the context of Laskowski inhibitor engineering, the way that various modifications intended to fine-tune an inhibitor's potency and selectivity impact on its association and dissociation rates remains unclear. This information is important as Laskowski inhibitors are becoming increasingly used as design templates to develop new protease inhibitors for pharmaceutical applications. In this study, we used the cyclic peptide, sunflower trypsin inhibitor-1 (SFTI-1), as a model system to explore how the inhibitor's sequence and structure relate to its binding kinetics and function. Using enzyme assays, MD simulations and NMR spectroscopy to study SFTI variants with diverse sequence and backbone modifications, we show that the geometry of the binding loop mainly influences the inhibitor's potency by modulating the association rate, such that variants lacking a favourable conformation show dramatic losses in activity. Additionally, we show that the inhibitor's sequence (including both the binding loop and its scaffolding) influences its potency and selectivity by modulating both the association and the dissociation rates. These findings provide new insights into protease inhibitor function and design that we apply by engineering novel inhibitors for classical serine proteases, trypsin and chymotrypsin and two kallikrein-related peptidases (KLK5 and KLK14) that are implicated in various cancers and skin diseases.
Keyword Cyclic peptides
Drug design
Kallikrein-related peptidase
Laskowski mechanism
Q-Index Code C1
Q-Index Status Confirmed Code
Institutional Status UQ

Document type: Journal Article
Sub-type: Article (original research)
Collections: Official 2016 Collection
School of Biomedical Sciences Publications
Institute for Molecular Bioscience - Publications
 
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