Cystatin Activity–Based Protease Profiling to Select Protease Inhibitors Useful in Plant Protection

No Thumbnail Available
File version
Author(s)
Goulet, MC
Sainsbury, F
Michaud, D
Griffith University Author(s)
Primary Supervisor
Other Supervisors
Editor(s)
Date
2020
Size
File type(s)
Location
License
Abstract

Protease inhibitors of the cystatin protein superfamily show potential in plant protection for the control of herbivorous pests. Here, we describe a cystatin activity–based profiling procedure for the selection of potent cystatin candidates, using single functional variants of tomato cystatin SlCYS8 and digestive Cys proteases of the herbivore insect Colorado potato beetle as a case study. The procedure involves the capture of target Cys proteases with biotinylated versions of the cystatins, followed by the identification and quantitation of captured proteases by mass spectrometry. An example is given to illustrate usefulness of the approach as an alternative to current procedures for recombinant inhibitor selection based on in vitro assays with synthetic peptide substrates. A second example is given showing its usefulness as a tool to compare the affinity spectra of inhibitor variants toward different subsets of target protease complements.

Journal Title

Methods in Molecular Biology

Conference Title
Book Title
Edition
Volume

2139

Issue
Thesis Type
Degree Program
School
Publisher link
Patent number
Funder(s)
Grant identifier(s)
Rights Statement
Rights Statement
Item Access Status
Note
Access the data
Related item(s)
Subject

Other chemical sciences

Biochemistry and cell biology

Biotinylated cystatins

Cys protease capture

Cystatin activity–based protease profiling

Herbivorous insect digestive proteases

Plant protease inhibitors

Persistent link to this record
Citation

Goulet, MC; Sainsbury, F; Michaud, D, Cystatin Activity–Based Protease Profiling to Select Protease Inhibitors Useful in Plant Protection, Methods in Molecular Biology, 2020, 2139, pp. 353-366

Collections