Cloning, expression, crystallization and preliminary X-ray analysis of the first two Ig domains from human Roundabout 1 (Robo1)

Abstract

Activation of Roundabout 1 (Robo1) by Slit proteins results in axon repulsion from the midline. Robo1 is a large transmembrane receptor expressed on the axon growth cone and the minimal Robo1-binding region required for Slit activation has been mapped to the N-terminal Ig1-2 domains. The cDNA encoding the first two Ig domains of Robo1 has been cloned and the protein has been expressed in HEK293 EBNA-1 mammalian cells. Here, the purification and crystallization conditions of this Robo1 construct are reported. The crystals are orthorhombic, space group P2(1)2(1)2, with unit-cell parameters a = 38.8, b = 69.4, c = 103.3 A and one molecule in the asymmetric unit. X-ray diffraction data have been collected to 2.8 A resolution on beamline ID29 at the ESRF.