Characterizing the N- and C-terminal SUMO Interacting Motifs of the Scaffold Protein DAXX

Abstract

DAXX is a scaffold protein with diverse roles that often depend upon binding SUMO via its N- and/or C-terminal SUMO-interacting motifs (SIM-N and SIM-C). In each case, binding is mediated by hydrophobic and electrostatic interactions, and weakens with increasing ionic strength. Neither isolated SIM shows any significant paralog specificity, and the measured μM-range KD values of SIM-N towards both SUMO-1 and SUMO-2 are ~4 fold lower than those of SIM-C. Furthermore, SIM-N binds SUMO-1 predominantly in a parallel orientation, whereas SIM-C interconverts between parallel and antiparallel binding modes on a msec-μsec timescale. The differences in affinities and binding modes are attributed to the differences in charged residues that flank the otherwise identical hydrophobic core sequences of the two SIMs. In addition, within its native context, SIM-N binds intramolecularly to the adjacent N-terminal helical bundle domain of DAXX, thus reducing its apparent affinity for SUMO. This behavior suggests a possible auto-regulatory mechanism for DAXX. The interaction of a C-terminal fragment of DAXX with an N-terminal fragment of the sumoylated Ets1 transcription factor is mediated by SIM-C. Importantly, this interaction does not involve any direct contacts between DAXX and Ets1, but rather derives from the non-covalent binding of SIM-C to SUMO-1, which in turn is covalently linked to the unstructured N-terminal segment of Ets1. These results provide insights into the binding mechanisms and hence biological roles of the DAXX SUMO-interacting motifs.

The article is published online in the Journal of Biological Chemistry and is free to access.