Fig. 1.

MORC2 is a GHKL-type ATPase. a Domain organization of human MORC2. The GHKL ATP binding domain and the transducer-like domain (trans.) together form the ATPase module, as marked. CC indicates a predicted coiled coil; CW indicates a CW-type zinc finger domain; TCD indicates a predicted tudor-chromodomain. b Fitted T_m s derived from differential scanning fluorimetry (DSF) for several MORC2 variants at 5 µM, in the absence (solid bar) and presence (striped bar) of 2 mM Mg-AMPPNP. This non-hydrolysable ATP analog significantly increases the thermal stability of wild-type (WT) MORC2(1–603), while the N39A point mutant abrogates binding and WT MORC2(1–282) is stabilized to a much smaller extent than WT MORC2(1–603). Quoted T_m values are an average of at least two replicates; note that the deviation between these measurements was <0.2 °C in all cases. c Portions of overlaid SEC-MALS UV traces for 40 µM WT MORC2(1–603) in the absence (solid line) and presence (dashed line) of 2 mM Mg-AMPPNP. The MALS data across the center of the major peaks in each case are shown on the right-hand axis, and are consistent with monomeric (expected mass: 70 kDa) and dimeric (expected mass: 140 kDa) species for the apo and AMPPNP-bound protein, respectively. Also shown are the fitted hydrodynamic radii obtained from QELS analysis of the peaks. The peak at 48 min in the AMPPNP-treated trace is the elution of excess (unbound) nucleotide. d Rate of ATP hydrolysis by wild-type (WT) and N39A MORC2(1–603) variants at 37 °C in the presence of 7.5 mM ATP, measured using an NADH-coupled continuous assay. Error bars represent standard deviation between measurements; n = 8 (WT), n = 7 (N39A). e Steady-state ATPase activity of 4 µM WT MORC2(1–603) at 37 °C fitted to a model of Michaelis–Menten kinetics