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A survey of furin substrate specificity using substrate phage display
D J Matthews1, L J Goodman, C M Gorman
1Department of Protein Engineering, Genentech, Inc., South San Francisco, California 94080.
Protein Science : a Publication of the Protein Society
|August 1, 1994
Summary
Researchers used substrate phage display to identify furin enzyme cleavage sites. They discovered a consensus RxxR motif and found that furin
Area of Science:
- Proteomics
- Enzymology
- Molecular Biology
Background:
- Furin is a mammalian enzyme crucial for cleaving protein precursors.
- Understanding furin's substrate specificity is key to studying protein processing.
Purpose of the Study:
- To elucidate the substrate specificity of the furin enzyme.
- To identify consensus substrate motifs recognized by furin.
Main Methods:
- Substrate phage display was employed to screen a library of potential substrates.
- Affinity chromatography was used to purify cleaved phage displaying furin substrates.
- DNA sequencing identified the enriched substrate sequences.
- Substrate-alkaline phosphatase fusion protein system validated cleavage sites.
Main Results:
- A prevalent RxxR motif was identified in cleaved substrates.
- Substrates with Pro or Thr at the P2 position showed efficient cleavage.
- A 7-residue motif (L/P)RRF(K/R)RP was suggested for extended furin recognition.
Conclusions:
- The study clarifies furin's substrate specificity, highlighting the RxxR motif and extended recognition.
- Substrate phage display is a powerful tool for identifying protease consensus motifs.