Cryo-EM structures of the MnmE-MnmG complex reveal large conformational changes and provide new insights into the mechanism of tRNA modification

Abstract

MnmE and MnmG form a conserved protein complex responsible for the addition of a 5-carboxymethylaminomethyl (cmnm5) group onto the wobble uridine of several tRNAs. Within this complex, both proteins collaborate intensively to catalyze a tRNA modification reaction that involves glycine as a substrate in addition to three different cofactors, with FAD and NADH binding to MnmG and methylenetetrahydrofolate (5,10-CH2-THF) to MnmE. Without structures of the MnmEG complex it remained enigmatic how these substrates and co-factors can be brought together in a concerted manner. Prior SAXS data suggested that the MnmE (2) and MnmG ({beta}2) homodimers can adopt either an 2{beta}2 or 4{beta}2 complex, depending on the nucleotide state of MnmE. Here, we report the cryo-EM structures of the MnmEG complex in the 2{beta}2 and 4{beta}2 oligomeric states. These structures reveal that MnmE undergoes large conformational changes upon interaction with MnmG, resulting in an asymmetric MnmE dimer. In particular, the functionally important C-terminal helix of MnmE relocates from the 5,10-CH2-THF binding pocket of MnmE to the FAD binding pocket of MnmG, thus suggesting a mechanism for the transfer of an activated methylene group from one active site to the other. Together, these findings provide crucial new insights into the MnmEG-catalyzed reaction. Graphical abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=89 SRC="FIGDIR/small/625835v1_ufig1.gif" ALT="Figure 1"> View larger version (26K): org.highwire.dtl.DTLVardef@144e41forg.highwire.dtl.DTLVardef@1dce537org.highwire.dtl.DTLVardef@14c9359org.highwire.dtl.DTLVardef@5dec04_HPS_FORMAT_FIGEXP M_FIG C_FIG