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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


IMPACT FACTOR 2018: 3.014
5-year IMPACT FACTOR: 3.162

CiteScore 2018: 3.09

SCImago Journal Rank (SJR) 2018: 1.482
Source Normalized Impact per Paper (SNIP) 2018: 0.820

Online
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1437-4315
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Volume 393, Issue 5

Issues

Non-combinatorial library screening reveals subsite cooperativity and identifies new high-efficiency substrates for kallikrein-related peptidase 14

Simon J. de Veer
  • Institute of Health and Biomedical Innovation, Queensland University of Technology, Brisbane, Queensland 4059, Australia
  • Other articles by this author:
  • De Gruyter OnlineGoogle Scholar
/ Joakim E. Swedberg
  • Institute for Molecular Bioscience, University of Queensland, Brisbane, Queensland 4072, Australia
  • Other articles by this author:
  • De Gruyter OnlineGoogle Scholar
/ Edward A. Parker
  • Institute of Health and Biomedical Innovation, Queensland University of Technology, Brisbane, Queensland 4059, Australia
  • Other articles by this author:
  • De Gruyter OnlineGoogle Scholar
/ Jonathan M. Harris
  • Institute of Health and Biomedical Innovation, Queensland University of Technology, Brisbane, Queensland 4059, Australia
  • Other articles by this author:
  • De Gruyter OnlineGoogle Scholar

Abstract

An array of substrates link the tryptic serine protease, kallikrein-related peptidase 14 (KLK14), to physiological functions including desquamation and activation of signaling molecules associated with inflammation and cancer. Recognition of protease cleavage sequences is driven by complementarity between exposed substrate motifs and the physicochemical signature of an enzyme’s active site cleft. However, conventional substrate screening methods have generated conflicting subsite profiles for KLK14. This study utilizes a recently developed screening technique, the sparse matrix library, to identify five novel high-efficiency sequences for KLK14. The optimal sequence, YASR, was cleaved with higher efficiency (kcat/Km=3.81±0.4×106 m-1 s-1) than favored substrates from positional scanning and phage display by 2- and 10-fold, respectively. Binding site cooperativity was prominent among preferred sequences, which enabled optimal interaction at all subsites as indicated by predictive modeling of KLK14/substrate complexes. These simulations constitute the first molecular dynamics analysis of KLK14 and offer a structural rationale for the divergent subsite preferences evident between KLK14 and closely related KLKs, KLK4 and KLK5. Collectively, these findings highlight the importance of binding site cooperativity in protease substrate recognition, which has implications for discovery of optimal substrates and engineering highly effective protease inhibitors.

Keywords: phage display; positional scanning synthetic combinatorial library; serine protease; sparse matrix library; substrate specificity

About the article

Corresponding author


Received: 2011-11-07

Accepted: 2011-12-05

Published in Print: 2012-05-01


Citation Information: , Volume 393, Issue 5, Pages 331–341, ISSN (Online) 1437-4315, ISSN (Print) 1431-6730, DOI: https://doi.org/10.1515/bc-2011-250.

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