Chronicles of a Demented Mind

Information on protein stability is essential for studying protein structure and folding and can also be used indirectly to monitor proteinligand or proteinprotein interactions. While clearly valuable, the experimental determination of a protein’s stability typically requires biophysical instrumentation and substantial quantities of purified protein, which has limited the use of this technique as a general laboratory method Cheap Nike Air Max shoes. We report here a simple new method for determining protein stability by using pulse proteolysis with varying concentrations of denaturant. Pulse proteolysis is designed to digest only the unfolded proteins in an equilibrium mixture of folded and unfolded proteins that relaxes on a time scale longer than the proteolytic pulse. We used this method to study the stabilities of Escherichia coli ribonuclease H and its variants, both in purified form and directly from cell lysates. The G

unf values obtained by this technique were in agreement with those determined by traditional methods. We also successfully used this method to monitor the binding of maltosebinding protein to maltose, as well as to rapidly screen cognate ligands for this protein Nike Air Max 2009 Womens. The simplicity of pulse proteolysis suggests that it is an excellent strategy for the highthroughput determination of protein stability in protein engineering and drug discovery applications Nike Air Max for sale.

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