Jump to ContentJump to Main Navigation
Show Summary Details

Buchner, Johannes

Biological Chemistry

Editor-in-Chief: Brüne, Bernhard

Editorial Board Member: Buchner, Johannes / Lei, Ming / Ludwig, Stephan / Sies, Helmut / Turk, Boris / Wittinghofer, Alfred

SCImago Journal Rank (SJR) 2015: 1.607
Source Normalized Impact per Paper (SNIP) 2015: 0.751
Impact per Publication (IPP) 2015: 2.609

See all formats and pricing



Select Volume and Issue


Identifi cation of protease exosite-interacting peptides that enhance substrate cleavage kinetics

Abeer M. Jabaiah1 / Jennifer A. Getz1 / Witold A. Witkowski2 / Jeanne A. Hardy2 / 1

1Department of Chemical Engineering, University of California, Santa Barbara, CA 93106, USA

2Department of Chemistry, University of Massachusetts, Amherst, 710 North Pleasant Street, Amherst, MA 01003-9336, USA

Corresponding author

Citation Information: . Volume 393, Issue 9, Pages 933–941, ISSN (Online) 1437-4315, ISSN (Print) 1431-6730, DOI: https://doi.org/10.1515/hsz-2012-0162, September 2012

Publication History


This article offers supplementary material which is provided at the end of the article.


Many peptidases are thought to require non-active site interaction surfaces, or exosites, to recognize and cleave physiological substrates with high specifi city and catalytic effi ciency. However, the existence and function of protease exosites remain obscure owing to a lack of effective methods to identify and characterize exosite-interacting substrates. To address this need, we modifi ed the cellular libraries of peptide substrates (CLiPS) methodology to enable the discovery of exosite-interacting peptide ligands. Invariant cleavage motifs recognized by the active sites of thrombin and caspase-7 were displayed on the outer surface of bacteria adjacent to a candidate exosite-interacting peptide. Exosite peptide libraries were then screened for ligands that accelerate cleavage of the active site recognition motif using two-color fl ow cytometry. Exosite CLiPS (eCLiPS) identifi ed exosite-binding peptides for thrombin that were highly similar to a critical exosite interaction motif in the thrombin substrate, proteaseactivated receptor 1. Protease activity probes incorporating exosite-binding peptides were cleaved ten-fold faster than substrates without exosite ligands, increasing their sensitivity to thrombin activity in vitro. For comparison, screening with caspase-7 yielded peptides that modestly enhanced (two-fold) substrate cleavage rates. The eCLiPS method provides a new tool to profi le the ligand specifi city of protease exosites and to develop improved substrates.

Keywords: CLiPS; exosite; protease; substrate; thrombin

Supplementary Article Materials

Citing Articles

Here you can find all Crossref-listed publications in which this article is cited. If you would like to receive automatic email messages as soon as this article is cited in other publications, simply activate the “Citation Alert” on the top of this page.

Magdalena Wysocka, Natalia Gruba, Renata Grzywa, Artur Giełdoń, Remigiusz Bąchor, Krzysztof Brzozowski, Marcin Sieńczyk, Jenne Dieter, Zbigniew Szewczuk, Krzysztof Rolka, and Adam Lesner
Scientific Reports, 2016, Volume 6, Page 22856
Shrikrishnan Sankaran, Mustafa Can Kiren, and Pascal Jonkheijm
ACS Nano, 2015, Volume 9, Number 4, Page 3579

Comments (0)

Please log in or register to comment.