-
Robert T Byrne,
Fiona Whelan,
Pierre Aller,
Louise E Bird,
Adam Dowle,
Carina M C Lobley,
Yamini Reddivari,
Joanne E Nettleship,
Raymond J Owens,
Alfred A Antson,
David G Waterman,
F Stfc,
Rutherford Appleton
[show abstract]
[hide abstract]
ABSTRACT: Uridine at position 34 of bacterial transfer RNAs is commonly modified to uridine-5-oxyacetic acid (cmo 5 U) to increase the decoding capacity. The protein CmoA is involved in the formation of cmo 5 U and was annotated as an S-adenosyl-l-methionine-dependent (SAM-dependent) methyltransferase on the basis of its sequence homology to other SAM-containing enzymes. However, both the crystal structure of Escherichia coli CmoA at 1.73 Å resolution and mass spectrometry demonstrate that it contains a novel cofactor, S-adenosyl-S-carboxymethyl-l-homocysteine (SCM-SAH), in which the donor methyl group is substituted by a carboxy-methyl group. The carboxyl moiety forms a salt-bridge interaction with Arg199 that is conserved in a large group of CmoA-related proteins but is not conserved in other SAM-containing enzymes. This raises the possibility that a number of enzymes that have previously been annotated as SAM-dependent are in fact SCM-SAH-dependent. Indeed, inspec-tion of electron density for one such enzyme with known X-ray structure, PDB entry 1im8, suggests that the active site contains SCM-SAH and not SAM.
Acta Crystallographica Section D Biological Crystallography 05/2013; · 12.62 Impact Factor
-
[show abstract]
[hide abstract]
ABSTRACT: Dihydrouridine synthases catalyse the reduction of uridine to dihydrouridine in the D-loop and variable loop of tRNA. The human dihydrouridine synthase HsDus2L has been implicated in the development of pulmonary carcinogenesis. Here, the purification, crystallization and preliminary X-ray characterization of the HsDus2L catalytic domain are reported. The crystals belonged to space group P2(1) and contained a single molecule of HsDus2L in the asymmetric unit. A complete data set was collected to 1.9 Å resolution using synchrotron radiation.
Acta Crystallographica Section F Structural Biology and Crystallization Communications 03/2012; 68(Pt 3):333-6. · 0.51 Impact Factor
-
[show abstract]
[hide abstract]
ABSTRACT: MTH1203, a β-CASP metallo-β-lactamase family nuclease from the archaeon Methanothermobacter thermautotrophicus, was identified as a putative nuclease that might contribute to RNA processing. The crystal structure of MTH1203 reveals that, in addition to the metallo-β-lactamase nuclease and the β-CASP domains, it contains two contiguous KH domains that are unique to MTH1203 and its orthologs. RNA-binding experiments indicate that MTH1203 preferentially binds U-rich sequences with a dissociation constant in the micromolar range. In vitro nuclease activity assays demonstrated that MTH1203 is a zinc-dependent nuclease. MTH1203 is also shown to be a dimer and, significantly, this dimerization enhances the nuclease activity. Transcription termination in archaea produces mRNA transcripts with U-rich 3' ends that could be degraded by MTH1203 considering its RNA-binding specificity. We hypothesize that this nuclease degrades mRNAs of proteins targeted for degradation and so regulates archaeal RNA turnover, possibly in concert with the exosome.
Structure 05/2011; 19(5):622-32. · 6.35 Impact Factor
-
[show abstract]
[hide abstract]
ABSTRACT: Post-transcriptional nucleoside modifications fine-tune the biophysical and biochemical properties of transfer RNA (tRNA) so that it is optimized for participation in cellular processes. Here we report the crystal structure of unmodified tRNA(Phe) from Escherichia coli at a resolution of 3 A. We show that in the absence of modifications the overall fold of the tRNA is essentially the same as that of mature tRNA. However, there are a number of significant structural differences, such as rearrangements in a triplet base pair and a widened angle between the acceptor and anticodon stems. Contrary to previous observations, the anticodon adopts the same conformation as seen in mature tRNA.
Nucleic Acids Research 03/2010; 38(12):4154-62. · 8.03 Impact Factor
-
[show abstract]
[hide abstract]
ABSTRACT: MTH909 is the Methanothermobacter thermautotrophicus orthologue of Saccharomyces cerevisiae TAN1, which is required for N(4)-acetylcytidine formation in tRNA. The protein consists of an N-terminal near-ferredoxin-like domain and a C-terminal THUMP domain. Unlike most other proteins containing the THUMP domain, TAN1 lacks any catalytic domains and has been proposed to form a complex with a catalytic protein that is capable of making base modifications. MTH909 has been cloned, overexpressed and purified. The molecule exists as a monomer in solution. X-ray data were collected to 2.85 A resolution from a native crystal belonging to space group P6(1)22 (or P6(5)22), with unit-cell parameters a = 69.9, c = 408.5 A.
Acta Crystallographica Section F Structural Biology and Crystallization Communications 12/2008; 64(Pt 11):1083-6. · 0.51 Impact Factor
