Related Experiment Video
Updated: May 24, 2026

A High-Throughput Luciferase Assay to Evaluate Proteolysis of the Single-Turnover Protease PCSK9
Published on: August 28, 2018
Active site mutations change the cleavage specificity of neprilysin
Travis Sexton1, Lisa J Hitchcook, David W Rodgers
1Department of Molecular and Cellular Biochemistry, The Center for Structural Biology, University of Kentucky, Lexington, Kentucky, United States of America.
Abstract:
Neprilysin (NEP), a member of the M13 subgroup of the zinc-dependent endopeptidase family is a membrane bound peptidase capable of cleaving a variety of physiological peptides. We have generated a series of neprilysin variants containing mutations at either one of two active site residues, Phe(563) and Ser(546). Among the mutants studied in detail we observed changes in their activity towards leucine(5)-enkephalin, insulin B chain, and amyloid β(1-40). For example, NEP(F563I) displayed an increase in preference towards cleaving leucine(5)-enkephalin relative to insulin B chain, while mutant NEP(S546E) was less discriminating than neprilysin. Mutants NEP(F563L) and NEP(S546E) exhibit different cleavage site preferences than neprilysin with insulin B chain and amyloid ß(1-40) as substrates. These data indicate that it is possible to alter the cleavage site specificity of neprilysin opening the way for the development of substrate specific or substrate exclusive forms of the enzyme with enhanced therapeutic potential.
More Related Videos
08:56Demonstration of Proteolytic Activation of the Epithelial Sodium Channel (ENaC) by Combining Current Measurements with Detection of Cleavage Fragments
Published on: July 5, 2014
11:49Generation of Centromere-Associated Protein-E CENP-E-/- Knockout Cell Lines using the CRISPR/Cas9 System
Published on: June 23, 2023
Related Concept Videos
Conservative Site-specific Recombination and Phase Variation
The recognition sites for Cre recombinase called LoxP...
Mutations
Ligand Binding and Linkage
Allosteric Proteins-ATCase
Aspartate transcarbamoylase (ATCase) is a cytosolic enzyme that catalyzes the condensation of L-aspartate and carbamoyl phosphate to N-carbamoyl-L-aspartate. This reaction is the first step in pyrimidine biosynthesis. UTP and CTP, the end products of the pyrimidine synthesis pathway,...
Caspases
RNA Editing