PMID:23150671

Citation	'Boundy, S, Safo, MK, Wang, L, Musayev, FN, O'Farrell, HC, Rife, JP and Archer, GL (2012) Characterization of the Staphylococcus aureus rRNA Methyltransferase Encoded by orfX, the Gene Containing the Staphylococcus aureus SCCmec Insertion Site. J. Biol. Chem. '
Abstract	The gene orfX is conserved among all staphylococci and its complete sequence is maintained upon insertion of the genomic island SCCmec, containing the gene encoding resistance to beta-lactam antibiotics (mecA), into its carboxy-terminus. The function of OrfX has not been determined. We showed that OrfX was constitutively produced during growth, that orfX could be inactivated without altering bacterial growth and that the insertion of SCCmec did not alter gene expression. We solved the crystal structure of OrfX at 1.7 Å and found that it belonged to the S-adenosyl methionine (AdoMet)-dependent α/β-knot superfamily of SPOUT methyltransferases (MTases), with a high structural homology to YbeA, the gene product of the E. coli 70S ribosomal MTase rlmH. MTase activity was confirmed by demonstrating the OrfX dependent methylation of the Staphylococcus aureus 70S ribosome. When OrfX was crystallized in the presence of its AdoMet substrate, it was found that each monomer of the homodimeric structure binds AdoMet in its active site. Solution studies using isothermal titration calorimetry confirmed that each monomer bound AdoMet but with different binding affinities (Kd values of 52 ±0.4 μM and of 606 ±2 μM). In addition, the structure showed that the AdoMet binding pocket, formed by a deep trefoil knot, contained a bound phosphate molecule, which was the likely nucleotide methylation site. This study represents the first characterization of a staphylococcal ribosomal MTase, and is the first crystal structure of a member of the α/β- knot superfamily of SPOUT MTases in the RlmH or COG1576 family with bound AdoMet.
Links	PubMed Online version:10.1074/jbc.M112.385138
Keywords
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Annotations

Gene product	Qualifier	GO Term	Evidence Code	with/from	Aspect	Extension	Notes	Status
STAAU:Q1HAH9		GO:0008649: rRNA methyltransferase activity	ECO:0000315:		F		fig 3	complete CACAO 5922
STAAU:Q1HAH9	enables	GO:0008649: rRNA methyltransferase activity	ECO:0000315: mutant phenotype evidence used in manual assertion		F		Seeded From UniProt	complete

References

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