Fig. 3. Interaction of SpoIIE, SpoIIE (II–III) and SpoIIE (III). (A) SDS–PAGE of SpoIIE (II–III) and SpoIIE (III) after separation on a Superdex 200 gel filtration column and staining with Coomassie Blue. SpoIIE (II–III) was eluted between 59 and 77 ml, corresponding to an apparent M_r between 300 and 50 kDa. SpoIIE (III) was eluted between 84 and 90 ml, corresponding to an apparent M_r of ∼28 kDa. The upper and lower arrows indicate the position of SpoIIE (II–III) and SpoIIE (III), respectively. (B) Purified full-length SpoIIE (3.2 μM) and SpoIIE (III) (3 μM) were mixed and applied to a Superdex 200 gel filtration column equilibrated with buffer C. Elution was performed in the same buffer and fractions of 1 ml were collected and monitored by Western blotting with anti-SpoIIE antibodies. Full-length SpoIIE was eluted between 46 and 66 ml, corresponding to an apparent M_r between 700 and 170 kDa. SpoIIE (III) was eluted between 84 and 91 ml, corresponding to an apparent M_r of ∼28 kDa. The positions of full-length SpoIIE and SpoIIE (III) are indicated by solid arrows. The additional immunoreactive bands (indicated by dotted lines) correspond to dimeric and higher molecular mass (possibly tetrameric) forms of SpoIIE.