Mapping Site-Specific Changes that Affect Stability of the NTerminal Domain of Calmodulin
Krause, Mary Elizabeth
Martin, Talia Thresa
Laurence, Jennifer S.
American Chemical Society
Scholarly/refereed, author accepted manuscript
This document is the Accepted Manuscript version of a Published Work that appeared in final form in Molecular Pharmaceutics, copyright © American Chemical Society after peer review and technical editing by the publisher. To access the final edited and published work see http://doi.org/10.1021/mp2004109.
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Biophysical tools have been invaluable in formulating therapeutic proteins. These tools characterize protein stability rapidly in a variety of solution conditions, but in general, the techniques lack the ability to discern site-specific information to probe how solution environment acts to stabilize or destabilize the protein. NMR spectroscopy can provide site-specific information about subtle structural changes of a protein under different conditions, enabling one to assess the mechanism of protein stabilization. In this study, NMR was employed to detect structural perturbations at individual residues as a result of altering pH and ionic strength. The N-terminal domain of calmodulin (N-CaM) was used as a model system, and the 1H-15N heteronuclear single quantum coherence (HSQC) experiment was used to investigate effects of pH and ionic strength on individual residues. NMR analysis revealed that different solution conditions affect individual residues differently, even when the amino acid sequence and structure are highly similar. This study shows that addition of NMR to the formulation toolbox has the ability to extend understanding of the relationship between site-specific changes and overall protein stability.
Krause, M. E., Martin, T. T., & Laurence, J. S. (2012). Mapping Site-Specific Changes that Affect Stability of the N-Terminal Domain of Calmodulin. Molecular Pharmaceutics, 9(4), 734–743. http://doi.org/10.1021/mp2004109
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