Fig. 2.

The R64P lamin protein makes only short filaments. (A) (a–d) WT Ce-lamin. (e–h) R64P mutant protein. The assembly reaction was stopped after 5 sec (a and e), 15 sec (b and f), 5 min (c and g), or overnight (d and h) by adding an equal amount of 0.2% (vol/vol) glutaraldehyde (17). Samples were negatively stained and visualized by electron microscopy. (Scale bars: 100 nm.) (B) R64P Ce-lamin was assembled overnight under standard conditions and assembly products were centrifuged at 11,500 × g. The supernatant was transferred into a new tube and centrifuged at 26,000 × g, and the procedure was repeated at 104,000 × g. The precipitants of each step and the supernatant that remained after the last centrifugation step (SN) were analyzed in PAGE/12% SDS.