Mapping protease substrates by using a biotinylated phage substrate library

Chembiochem : a European Journal of Chemical Biology
Michael D ScholleBrian K Kay

Abstract

We describe a bacteriophage M13 substrate library encoding the AviTag (BirA substrate) and combinatorial heptamer peptides displayed at the N terminus of the mature form of capsid protein III. Phages are biotinylated efficiently (> or = 50%) when grown in E. coli cells coexpressing BirA, and such viral particles can be immobilized on a streptavidin-coated support and released by protease cleavage within the combinatorial peptide. We have used this library to map the specificity of human Factor Xa and a neuropeptidase, neurolysin (EC3.4.24.16). Validation by analysis of isolated peptide substrates has revealed that neurolysin recognizes the motif hydrophobic-X-Pro-Arg-hydrophobic, where Arg-hydrophobic is the scissile bond.

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Citations

Apr 3, 2007·Applied Microbiology and Biotechnology·Tetsuya KadonosonoMitsuyoshi Ueda
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Oct 19, 2017·Nature Communications·Michael S PackerDavid R Liu
Sep 26, 2020·Proceedings of the National Academy of Sciences of the United States of America·Jie ZhouJames A Wells
Nov 11, 2020·ACS Combinatorial Science·Sabrina E IskandarAlbert A Bowers

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