Changes in biophysical characteristics of PFN1 due to mutation causing amyotrophic lateral sclerosis.

Single amino acid mutations in profilin 1 (PFN1) have been found to cause amyotrophic lateral sclerosis (ALS). Recently, we developed a mouse model for ALS using a PFN1 mutation (glycine 118 to valine, G118V), and we are now interested in understanding how PFN1 becomes toxically lethal with only one amino acid substitution. Therefore, we studied mutation-related changes in the PFN1 protein and hypothesized that such changes significantly disturb its structure. Initially, we expressed and studied the purified PFN1 and PFN1 proteins from bacterial culture. We found that the PFN1 protein has a different mean residue ellipticity, as measured by far-UV circular dichroism, accompanied by a spectral shift. The intrinsic fluorescence of PFN1 showed a small fluctuation in maximum fluorescence absorption and intensity. Moreover, we examined the time course of PFN1 aggregation using SDS-PAGE, western blotting, and MALDI-TOF/TOF and found that compared with PFN1, PFN1 had an increased tendency to form aggregates. Dynamic light scattering data confirmed this, showing a larger size distribution for PFN1. Our data explain why PFN1 tends to aggregate, a phenotype that may be the basis for its neurotoxicity.