Mapping protease substrates using a biotinylated phage substrate library.
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.
- Research Organization:
- Argonne National Lab. (ANL), Argonne, IL (United States)
- Sponsoring Organization:
- USDOE Laboratory Directed Research and Development (LDRD) Program
- DOE Contract Number:
- DE-AC02-06CH11357
- OSTI ID:
- 932921
- Report Number(s):
- ANL/BIO/JA-54719; TRN: US200814%%178
- Journal Information:
- ChemBioChem, Vol. 7, Issue 5 ; May 5, 2005
- Country of Publication:
- United States
- Language:
- ENGLISH
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