Cloning, expression, crystallization and preliminary X-ray analysis of the first two Ig domains from human roundabout 1 (Robo1).
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.
European Molecular Biology Laboratory, 6 Rue Jules Horowitz, Grenoble, France.
Cecile
C
Morlot
Wieger
W
Hemrika
Roland A
RA
Romijn
Piet
P
Gros
Stephen
S
Cusack
Andrew A
AA
McCarthy
D002460:Cell Line; D003001:Cloning, Molecular; D003460:Crystallization; D018360:Crystallography, X-Ray; D006023:Glycoproteins / Q000737:chemistry / Q000378:metabolism; D006801:Humans; D007136:Immunoglobulins / Q000096:biosynthesis / Q000737:chemistry* / Q000235:genetics*; D009419:Nerve Tissue Proteins / Q000096:biosynthesis / Q000737:chemistry* / Q000235:genetics* / Q000378:metabolism; D010449:Peptide Mapping; D017434:Protein Structure, Tertiary; D011971:Receptors, Immunologic / Q000096:biosynthesis / Q000737:chemistry* / Q000235:genetics*; D000097905:Roundabout Proteins
D006023:Glycoproteins; D007136:Immunoglobulins; D009419:Nerve Tissue Proteins; D011971:Receptors, Immunologic; D000097905:Roundabout Proteins; C120566:slit protein, vertebrate