Structural Organization of the Nine Spectrin Repeats of Kalirin

Sequence analysis suggests that KALRN, a Rho GDP/GTP exchange factor genetically linked to schizophrenia, could contain as many as nine tandem spectrin repeats (SRs). We expressed and purified fragments of Kalirin containing from one to five putative SRs to determine whether they formed nested structures that could endow Kalirin with the flexible rodlike properties characteristic of spectrin and dystrophin. Far-UV circular dichroism studies indicated that Kalirin contains nine SRs. On the basis of thermal denaturation, sensitivity to chemical denaturants, and the solubility of pairs of repeats, the nine SRs of Kalirin form nested structures. Modeling studies confirmed this conclusion and identified an exposed loop in SRS; consistent with the modeling, this loop was extremely labile to proteolytic cleavage. Analysis of a direpeat fragment (SR4:5) encompassing the region of Kalirin known to interact with NOS2, DISC-1, PAM, and Arf6 identified this as the least stable region. Analytical ultracentrifugation indicated that SR1:3, SR4:6, and SR7:9 were monomers and adopted an extended conformation. Gel filtration suggested that Delta Kal7, a natural isoform that includes SR5:9, was monomeric and was not more extended than SR5:9. Similarly, the nine SRs of Kal7, which was also monomeric, were not more extended than SR5:9. The rigidity and flexibility of the nine SRs of Kal7, which separate its essential N-terminal Sec14p domain from its catalytic domain, play an essential role in its contribution to the formation and function of dendritic spines.