Figure 4.

Semi-denaturing agarose gel electrophoresis on polymers formed in vitro. Purified Nab3¹³⁴, the L800A mutant derivative of Nab3¹³⁴, purified Nrd1⁶⁰ (unpolymerized monomer and polymerized), thioredoxin-Nrd1⁶⁰ fusion, and thioredoxin (TRX) were incubated with SDS at 22° (-) or 100°C (+), subjected to electrophoresis, and transferred to nitrocellulose. Membranes were probed with anti-S-tag antibody and developed with chemiluminescent reagents. The positions of monomeric (arrowheads) and polymeric (vertical bar) versions of the proteins are indicated, as are the size (kD) and migration positions of Precision Plus Protein Kaleidoscope molecular weight standards (Bio-Rad, Inc.). TRX and Nab3¹³⁴ samples from the composite panel on the left were run on different ends of the same gel. Note that Nrd1⁶⁰ monomers were not detectable but the monomeric TRX-Nrd1⁶⁰ fusion protein was.