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Aurora-A site specificity: a study with synthetic peptide substrates.
Stefano Ferrari1, Oriano Marin, Mario A Pagano
1Institute of Molecular Cancer Research, University of Zurich, Winterthurerstr. 190, CH-8057 Zurich, Switzerland.
The Biochemical Journal
|August 9, 2005
Summary
Aurora-A (AurA) is a basophilic kinase. Researchers identified its substrate specificity consensus sequence, R/K/N-R-X-S/T-B, aiding in predicting AurA targets and differentiating it from PKA.
Area of Science:
- Biochemistry
- Molecular Biology
- Cell Biology
Background:
- Aurora-A (AurA) is a protein kinase crucial for mitotic entry.
- Overexpression of AurA can lead to cellular transformation.
- Understanding AurA's substrate specificity is key to elucidating its functions.
Purpose of the Study:
- To determine the structural features governing Aurora-A substrate specificity.
- To identify the consensus phosphorylation motif for Aurora-A.
- To develop tools for distinguishing Aurora-A activity from other kinases like PKA.
Main Methods:
- Assaying human recombinant Aurora-A on various peptide substrates, including Kemptide derivatives.
- Synthesizing and testing peptides representing Aurora-A autophosphorylation sites.
- Comparing Aurora-A substrate preference with Protein Kinase A (PKA).
Main Results:
- Aurora-A identified as a basophilic Ser/Thr kinase with consensus R/K/N-R-X-S/T-B.
- Proline at the n+1 position abrogates AurA phosphorylation.
- A specific peptide (ALKRASLGAA) differentiates AurA from PKA.
- AurA autophosphorylates at Thr288 via an intermolecular mechanism, independent of PKA in vivo.
Conclusions:
- The study defines the substrate specificity consensus for Aurora-A.
- A novel peptide assay allows for differential screening of AurA and PKA.
- The findings provide a rational basis for predicting physiological substrates of Aurora-A.