Expression, purification, and characterization of recombinant 8 kDa gelsolin fragment.

A mutation (D187N/Y) in human plasma gelsolin (GSN) leads to the generation of an 8 kDa GSN fragment (8 kDa-GSN), and consequently causes the familial amyloidosis of Finnish type. Because of its faster kinetics of amyloid formation under physiologically relevant conditions, 8 kDa-GSN is used to explore gelsolin amyloidosis and screen small molecules that can disaggregate amyloids. However, the synthetic 8 kDa-GSN is expensive, and substantial quantities of 8 kDa-GSN are needed for the screen. Here we report a study to obtain recombinant 8 kDa-GSN with high yield from Escherichia coli. Firstly, 8 kDa-GSN in fusion with Mxe GyrA intein was purified by Ni-affinity chromatography. Then 8 kDa-GSN was released by intein-mediated protein cleavage, and separated from intein by ion-exchange chromatography. The yield of 8 kDa-GSN was only 1.5 mg/L from bacterial culture in the previous report, while it was improved to 4.25 mg/L in our study. Finally, the amyloidogenic property of 8 kDa-GSN was validated by circular dichroism spectrometry and dynamic light scattering.