TY - JOUR
T1 - Strong Anion Exchange for Studying Protein-DNA Interactions by H/D Exchange Mass Spectrometry
AU - Sperry, Justin B.
AU - Wilcox, J. Micah
AU - Gross, Michael L.
N1 - Funding Information:
These studies were supported by the National Institutes of Health and National Center for Research Resources (Grant P41RR000954). We thank S. Akashi and Y. Nishimura, Yokohama City University, Japan, for a generous gift of the human TRF2 DNA-binding domain protein.
PY - 2008/6
Y1 - 2008/6
N2 - The use of mass spectrometry to study protein-ligand interactions is expanding into more complex systems including protein-DNA interactions. The excess amount of a model DNA or, more typically, an oligodeoxynucleotide (ODN), needed to study such interactions in an amide hydrogen-deuterium (H/D) exchange experiment, for example, causes serious signal suppression in the protein analysis. We describe here a modification of the traditional H/D exchange protocol whereby we utilize a strong anion exchange column to rapidly remove the ODN from solution before MS analysis. We showed the successful incorporation of such a column in a study of two protein-ODN systems: (1) the DNA-binding domain of human telomeric repeat binding factor 2 with a telomeric oligodeoxynucleotide and (2) thrombin with the thrombin-binding aptamer. The approach gave no appreciable difference in back-exchange compared to a method in which no strong anion exchange (SAX) is used.
AB - The use of mass spectrometry to study protein-ligand interactions is expanding into more complex systems including protein-DNA interactions. The excess amount of a model DNA or, more typically, an oligodeoxynucleotide (ODN), needed to study such interactions in an amide hydrogen-deuterium (H/D) exchange experiment, for example, causes serious signal suppression in the protein analysis. We describe here a modification of the traditional H/D exchange protocol whereby we utilize a strong anion exchange column to rapidly remove the ODN from solution before MS analysis. We showed the successful incorporation of such a column in a study of two protein-ODN systems: (1) the DNA-binding domain of human telomeric repeat binding factor 2 with a telomeric oligodeoxynucleotide and (2) thrombin with the thrombin-binding aptamer. The approach gave no appreciable difference in back-exchange compared to a method in which no strong anion exchange (SAX) is used.
UR - https://www.scopus.com/pages/publications/43549090144
U2 - 10.1016/j.jasms.2008.03.003
DO - 10.1016/j.jasms.2008.03.003
M3 - Article
C2 - 18417359
AN - SCOPUS:43549090144
SN - 1044-0305
VL - 19
SP - 887
EP - 890
JO - Journal of the American Society for Mass Spectrometry
JF - Journal of the American Society for Mass Spectrometry
IS - 6
ER -