Hierarchical binding of cofactors to the AAA ATPase p97.

Rudolf Virchow Center for Experimental Biomedicine, University of Wuerzburg, Josef-Schneider-Str. 2, 97080 Wuerzburg, Germany. petra.haenzelmann@virchow.uni-wuerzburg.de
The hexameric AAA ATPase p97 is involved in several human proteinopathies and mediates ubiquitin-dependent protein degradation among other essential cellular processes. Via its N-terminal domain (N domain), p97 interacts with multiple regulatory cofactors including the UFD1/NPL4 heterodimer and members of the "ubiquitin regulatory X" (UBX) domain protein family; however, the principles governing cofactor selectivity remain to be deciphered. Our crystal structure of the FAS-associated factor 1 (FAF1)UBX domain in complex with the p97N domain reveals that the signature Phe-Pro-Arg motif known to be crucial for interactions of UBX domains with p97 adopts a cis-proline configuration, in contrast to a cis-trans mixture we derive for the isolated FAF1UBX domain. Biochemical studies confirm that binding critically depends on a proline at this position. Furthermore, we observe that the UBX proteins FAF1 and UBXD7 only bind to p97-UFD1/NPL4, but not free p97, thus demonstrating for the first time a hierarchy in p97-cofactor interactions.
Mesh Terms:
Adaptor Proteins, Signal Transducing, Adenosine Triphosphatases, Amino Acid Motifs, Amino Acid Sequence, Calorimetry, Carrier Proteins, Chromatography, Gel, Coenzymes, Conserved Sequence, Crystallography, X-Ray, Humans, Molecular Sequence Data, Nuclear Proteins, Protein Binding, Protein Interaction Domains and Motifs, Protein Structure, Quaternary, Protein Structure, Tertiary, Proteins, Surface Properties, Titrimetry, Ubiquitin, Ubiquitinated Proteins
Structure Jun. 08, 2011; 19(6);833-43 [PUBMED:21645854]
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