title: Structural basis for hypermodification of the wobble uridine in tRNA by bifunctional enzyme MnmC.
abstract: BACKGROUND
  Methylaminomethyl modification of uridine or 2-thiouridine (mnm5U34 or mnm5s2U34) at the wobble position of tRNAs specific for glutamate, lysine and arginine are observed in Escherichia coli and allow for specific recognition of codons ending in A or G. In the biosynthetic pathway responsible for this post-transcriptional modification, the bifunctional enzyme MnmC catalyzes the conversion of its hypermodified substrate carboxymethylaminomethyl uridine (cmnm5U34) to mnm5U34. MnmC catalyzes the flavin adenine dinucleotide (FAD)-dependent oxidative cleavage of carboxymethyl group from cmnm5U34 via an imine intermediate to generate aminomethyl uridine (nm5U34), which is subsequently methylated by S-adenosyl-L-methionine (SAM) to yield methylaminomethyl uridine (mnm5U34).
  
  
  RESULTS
  The X-ray crystal structures of SAM/FAD-bound bifunctional MnmC from Escherichia coli and Yersinia pestis, and FAD-bound bifunctional MnmC from Yersinia pestis were determined and the catalytic functions verified in an in vitro assay.
  
  
  CONCLUSIONS
  The crystal structures of MnmC from two Gram negative bacteria reveal the overall architecture of the enzyme and the relative disposition of the two independent catalytic domains: a Rossmann-fold domain containing the SAM binding site and an FAD containing domain structurally homologous to glycine oxidase from Bacillus subtilis. The structures of MnmC also reveal the detailed atomic interactions at the interdomain interface and provide spatial restraints relevant to the overall catalytic mechanism.
authors.affiliation: Albert Einstein College of Medicine, 1300 Morris Park Avenue, Bronx, New York 10461, USA. jukim@aecom.yu.edu
authors.forename: Jungwook
authors.initials: J
authors.lastname: Kim
authors.forename: Steven C
authors.initials: SC
authors.lastname: Almo
mesh_terms: D000595:Amino Acid Sequence; D001665:Binding Sites; D020134:Catalytic Domain; D018360:Crystallography, X-Ray; D004926:Escherichia coli / Q000201:enzymology / Q000235:genetics; D029968:Escherichia coli Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D005182:Flavin-Adenine Dinucleotide / Q000378:metabolism; D008969:Molecular Sequence Data; D009097:Multienzyme Complexes / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D012343:RNA, Transfer / Q000737:chemistry / Q000378:metabolism*; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000737:chemistry / Q000235:genetics; D012436:S-Adenosylmethionine / Q000737:chemistry / Q000378:metabolism; D016415:Sequence Alignment; D055672:Static Electricity; D013891:Thiouridine / Q000031:analogs & derivatives / Q000737:chemistry / Q000378:metabolism; D014529:Uridine / Q000737:chemistry / Q000378:metabolism*; D015010:Yersinia pestis / Q000201:enzymology / Q000378:metabolism
chemical_list: C105273:2-thiouridine; D029968:Escherichia coli Proteins; C488767:MnmC protein, E coli; D009097:Multienzyme Complexes; D011994:Recombinant Proteins; D013891:Thiouridine; D005182:Flavin-Adenine Dinucleotide; D012436:S-Adenosylmethionine; D012343:RNA, Transfer; D014529:Uridine