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Biological Chemistry

Editor-in-Chief: Brüne, Bernhard

Editorial Board: Buchner, Johannes / Lei, Ming / Ludwig, Stephan / Thomas, Douglas D. / Turk, Boris / Wittinghofer, Alfred


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Volume 397, Issue 12

Issues

Exploring the active site binding specificity of kallikrein-related peptidase 5 (KLK5) guides the design of new peptide substrates and inhibitors

Simon J. de Veer
  • Institute of Health and Biomedical Innovation, Queensland University of Technology, Kelvin Grove, QLD 4059, Brisbane, Australia
  • Other articles by this author:
  • De Gruyter OnlineGoogle Scholar
/ Joakim E. Swedberg
  • Institute for Molecular Bioscience, The University of Queensland, Brisbane, QLD 4072, Australia
  • Other articles by this author:
  • De Gruyter OnlineGoogle Scholar
/ Maria Brattsand / Judith A. Clements
  • Institute of Health and Biomedical Innovation, Queensland University of Technology, Kelvin Grove, QLD 4059, Brisbane, Australia
  • Australian Prostate Cancer Research Centre Queensland, Translational Research Institute, Brisbane, QLD 4102, Australia
  • Other articles by this author:
  • De Gruyter OnlineGoogle Scholar
/ Jonathan M. Harris
  • Corresponding author
  • Institute of Health and Biomedical Innovation, Queensland University of Technology, 60 Musk Avenue, Kelvin Grove, Brisbane, QLD 4059, Australia
  • Email
  • Other articles by this author:
  • De Gruyter OnlineGoogle Scholar
Published Online: 2016-02-19 | DOI: https://doi.org/10.1515/hsz-2016-0112

Abstract

Kallikrein-related peptidase 5 (KLK5) is a promising therapeutic target in several skin diseases, including Netherton syndrome, and is emerging as a potential target in various cancers. In this study, we used a sparse matrix library of 125 individually synthesized peptide substrates to characterize the binding specificity of KLK5. The sequences most favored by KLK5 were GRSR, YRSR and GRNR, and we identified sequence-specific interactions involving the peptide N-terminus by analyzing kinetic constants (kcat and KM) and performing molecular dynamics simulations. KLK5 inhibitors were subsequently engineered by substituting substrate sequences into the binding loop (P1, P2 and P4 residues) of sunflower trypsin inhibitor-1 (SFTI-1). These inhibitors were effective against KLK5 but showed limited selectivity, and performing a further substitution at P2′ led to the design of a new variant that displayed improved activity against KLK5 (Ki=4.2±0.2 nm), weak activity against KLK7 and 12-fold selectivity over KLK14. Collectively, these findings provide new insight into the design of highly favored binding sequences for KLK5 and reveal several opportunities for modulating inhibitor selectivity over closely related proteases that will be useful for future studies aiming to develop therapeutic molecules targeting KLK5.

Keywords: inhibitor engineering; peptide; serine protease; sparse matrix library; sunflower trypsin inhibitor-1

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About the article

aPresent address: Institute for Molecular Bioscience, The University of Queensland, Brisbane, QLD 4072, Australia.


Received: 2016-01-13

Accepted: 2016-02-16

Published Online: 2016-02-19

Published in Print: 2016-12-01


Citation Information: Biological Chemistry, Volume 397, Issue 12, Pages 1237–1249, ISSN (Online) 1437-4315, ISSN (Print) 1431-6730, DOI: https://doi.org/10.1515/hsz-2016-0112.

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