High Resolution Structural Characterization of the Human Spastin Protein and its Complex With Tubulin

Abstract

Hereditary spastic paraplegia (HSP) is a motor neuron disease caused by a progressive degeneration of the motor axons of the corticospinal tract. A number of different proteins have been implicated in the pathology of HSP, including mitochondrial proteins, a kinesin motor, spartin and spastin. Of the autosomal dominant cases of HSP, approximately 40% are caused by point mutations or exon deletions in spastin. Because of spastin's significant role in the development of HSP, there have been an increasing number of studies on this protein and its role in axonal degeneration. Recent studies have shown that spastin has a microtubule severing activity, and that this activity is linked to the manifestation of neurological disorders in Drosophila. Until recently very little structural information has been available for the spastin protein, and the mechanism by which it severs microtubules remains elusive. The goal of this study is to determine the high resolution crystal structure of the human spastin protein both alone and in complex with a domain of tubulin. To this end, we have expressed and purified a construct of human spastin and a C-terminal tubulin construct, to which spastin binds. We have also crystallized spastin and collected a 3.3 angstrom data set and are now attempting to crystallize the spastin-tubulin complex. Visualization of such a complex will provide details as to how spastin binds to and severs microtubules, and will also help to explain how point mutations in the spastin gene lead to altered protein activity and a disease phenotype.