Characterization of a multi-functional metal-mediated nuclease by MALDI-TOF mass spectrometry
U Puapaiboon1, J Jai-Nhuknan, J A Cowan
1Evans Laboratory of Chemistry, The Ohio State University, 100 West 18th Avenue, Columbus, OH 43210, USA.
Abstract:
Mass spectrometric analysis of reaction products allows simultaneous characterization of activities mediated by multifunctional enzymes. By use of MALDI-TOF mass spectrometry, the relative influence of magnesium and manganese promoted exonuclease and phosphatase activities of Esherichia coli exonuclease III have been quantitatively measured, offering a rapid and sensitive alternative to radioactivity quantification and gel electrophoresis procedures for determination of reaction rate constants. Manganese is found to promote higher levels of exonuclease activity, which could be a source of mutagenic effects if this ion were selected as the natural cofactor. Several potential applications of these methods to quantitative studies of DNA repair chemistry are also described.
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