Results and Discussion
Sup35 was consistently eluted in a stepwise manner. Two peaks were observed from non-denatured Ni2+ affinity chromatography (Fig 1A). Peak I was dissociated into three main bands on SDS-PAGE (Fig 1B), the Sup35 monomer band, an undetermined band (approximately 45 kDa, named as X1), and an outer membrane protein band (Omp F) [26]. The assignment of Omp F is based on its molecular weight shown on SDS-PAGE, the intensity of the band and the mass spectrum result (Table S1). The dissociation of peak I on SDS-PAGE had many continuous bands between the monomer and X1, which were defined as zone X2 (Fig 1B). Peak II exhibited a monomer band on SDS-PAGE with an intensity less than that under DTT. This indicates that part of the species in peak II is aggregated (S-aggregate) and caused by inter-Sup35 disulfide bonds. No intensity change could be observed for the protein bands associated with peak I on SDS-PAGE with the addition of DTT, thus excluding the formation of inter-Sup35 disulfide bonds.