CHANGE OF THERMAL-STABILITY OF COLICIN E7 TRIGGERED BY ACIDIC PH SUGGESTS THE EXISTENCE OF UNFOLDED INTERMEDIATE DURING THE MEMBRANE-TRANSLOCATION PHASE
Kf. Chak et al., CHANGE OF THERMAL-STABILITY OF COLICIN E7 TRIGGERED BY ACIDIC PH SUGGESTS THE EXISTENCE OF UNFOLDED INTERMEDIATE DURING THE MEMBRANE-TRANSLOCATION PHASE, Proteins, 32(1), 1998, pp. 17-25
Purified colicin E7 was analyzed by CD spectrum and gel filtration chr
omatography in a mimicking membrane-translocation phase. It was found
that the CD spectra of colicin E7 at pH 7 and pH 2.5 were similar. Alt
hough the melting temperature of the protein shifted from 54.5 degrees
C to 34 degrees C at low pH, the thermal denaturation curves of colic
in E7 at different pH conditions still fit a two-state model. These ex
perimental results imply that a minor structural change, triggered by
acidic pH, for instance, may reduce the energy required for protein me
lting. In contrast to the minor change in secondary structure at diffe
rent pH conditions, we observed that, in vitro, all monomeric colicin
E7s converted into multimer-like conformations after recovering from t
he partial unfolding process. This multimeric form of colicin can only
be dissociated by formamide and guanidine hydrochloride, indicating t
hat this protein complex is indeed formed by aggregation of the monome
ric colicins, Most interestingly, the aggregated colicins still perfor
m in vivo bacteriocidal activity. We suggest that in a partial unfoldi
ng state the colicin is prepared for binding to the specific targets f
or translocation through the membrane, However, in the absence of spec
ific targets in vitro these unfold intermediates may therefore aggrega
te into the multimeric form of colicins. Proteins 32:17-25, 1998, (C)
1998 Wiley-Liss, Inc.