SUBSTRATE COORDINATION IN RADICAL SAM DEHYDROGENASES

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This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. Primary support for the subproject and the subproject's principal investigator may have been provided by other sources, including other NIH sources. The Total Cost listed for the subproject likely represents the estimated amount of Center infrastructure utilized by the subproject, not direct funding provided by the NCRR grant to the subproject or subproject staff. Radical SAM dehydrogenases use the oxidizing power of a 5'-deoxyadenosyl radical (5'-dA?) to carry out two-electron oxidations of alcohols or thiols via intermediates containing unpaired electrons. The 5'-dA? initiates catalysis by abstracting a key hydrogen atom on the substrate. Recently, characterization of AtsB, a radical SAM enzyme that catalyzes the oxidation of a seryl residue on a cognate protein to a formylglycyl (fGly) residue, showed that it contained three [4Fe?4S] clusters, and a working model was put forth that suggested that one of the clusters binds in contact with the substrate to facilitate loss of the second electron from a substrate radical intermediate by inner-sphere electron transfer. This study aims to test this model by using an analog of the substrate in which the relevant seryl residue is replaced by a selenocysteinyl residue and analyzing for the interaction by Se XAS.