The denaturation of circular enterocin AS-48 by urea and guanidinium hydrochloride

The unfolding thermodynamics of the circular enterocin protein AS-48, produced by Enterococcus faecalis, has been studied. The native structure of the 70-amino-acid- long protein turned out to be extremely stable against heat and denaturant-induced unfolding. At pH 2.5 and low ionic strength, it denatures at 102 degreesC, while at 25 degreesC, the structure only unfolds in 6.3 M guanidiniurn hydrochloride (GuHCl) and does not unfold even in 8 M urea. A comparison of its thermal unfolding in water and in the presence of urea shows a good correspondence between the two DeltaG(w)(298) values, which are about 30 U mol(-1) at pH 2.5 and low ionic strength. The stability of the structure is highly dependent upon ionic strength and so GuHCl acts both as a denaturant and a stabilising agent. This seems to be why the DeltaG(w)(298) value calculated from the unfolding data in GuHCl is twice as high as in the absence of this salt. At least part of the high stability of native AS-48 can almost certainly be put down to its circular organization since other structural features are quite normal for a protein of this size. (C) 2002 Elsevier Science B.V. All rights reserved.