Heterologous overexpression of Sup35 in Escherichia coli leads to both monomer and complex states

Abstract The heterologous overexpression states of prion proteins play a critical role in understanding the mechanisms of prion‐related diseases. We report herein the identification of soluble monomer and complex states for a bakers' yeast prion, Sup35, when expressed in Escherichia coli . Two peaks are apparent with the elution of His‐tagged Sup35 by imidazole from a Ni 2+ affinity column. Peak I contains Sup35 in both monomer and aggregated states. Sup35 aggregate is abbreviated as C‐aggregate and includes a non‐fibril complex comprising Sup35 aggregate‐HSP90‐Dna K, ATP synthase β unit (chain D), 30S ribosome subunit, and Omp F. The purified monomer and C‐aggregate can remain stable for an extended period of time. Peak II contains Sup35 also in both monomer and aggregated (abbreviated as S‐aggregate) states, but the aggregated states are caused by the formation of inter‐Sup35 disulfide bonds. This study demonstrates that further assembly of Sup35 non‐fibril C‐aggregate can be interrupted by the chaperone repertoire system in E. coli .