Modification of DNA by a viral enzyme and charged tRNA

Abstract

AbstractBacteriophage enzymes synthesize varied and complex DNA hypermodifications. The enzyme encoded by the phage Mu genemomis necessary for post-replicative carbamoylmethyl addition to the exocyclic amine of deoxyadenosine in DNA during the lytic phase of the viral life-cycle. The molecular details of this modification reaction, including the molecular origins of the modification itself, have long eluded understanding. Here, we demonstrate that Mom co-opts the translational machinery of the host by harvesting activated glycine from charged tRNAGlyto hypermodify adenine. Based on this insight, we report the firstin vitroreconstitution of the Mu hypermodification from purified components. Using isotope labeling, we demonstrate that the carbamoyl nitrogen of the Mom modification is derived from theN6 of adenine, indicating an on-base rearrangement of theN6 aminoacylation product, possibly via a cyclic intermediate. Informed by the X-ray crystal structure of Mom, we have probed the location of the active site, identified a novel insertion, and established substrate specificities of the Mom enzyme.