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Small methyltransferase RlmH assembles a composite active site to methylate a ribosomal pseudouridine.

ABSTRACT: Eubacterial ribosomal large-subunit methyltransferase H (RlmH) methylates 23S ribosomal RNA pseudouridine 1915 (?1915), which lies near the ribosomal decoding center. The smallest member of the SPOUT superfamily of methyltransferases, RlmH lacks the RNA recognition domain found in larger methyltransferases. The catalytic mechanism of RlmH enzyme is unknown. Here, we describe the structures of RlmH bound to S-adenosyl-methionine (SAM) and the methyltransferase inhibitor sinefungin. Our structural and biochemical studies reveal catalytically essential residues in the dimer-mediated asymmetrical active site. One monomer provides the SAM-binding site, whereas the conserved C-terminal tail of the second monomer provides residues essential for catalysis. Our findings elucidate the mechanism by which a small protein dimer assembles a functionally asymmetric architecture.

SUBMITTER: Koh CS

PROVIDER: S-EPMC5430550 | biostudies-literature | 2017 Apr

REPOSITORIES: biostudies-literature

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Small methyltransferase RlmH assembles a composite active site to methylate a ribosomal pseudouridine.

Koh Cha San CS Madireddy Rohini R Beane Timothy J TJ Zamore Phillip D PD Korostelev Andrei A AA

Scientific reports 20170420 1

Eubacterial ribosomal large-subunit methyltransferase H (RlmH) methylates 23S ribosomal RNA pseudouridine 1915 (Ψ1915), which lies near the ribosomal decoding center. The smallest member of the SPOUT superfamily of methyltransferases, RlmH lacks the RNA recognition domain found in larger methyltransferases. The catalytic mechanism of RlmH enzyme is unknown. Here, we describe the structures of RlmH bound to S-adenosyl-methionine (SAM) and the methyltransferase inhibitor sinefungin. Our structural ...[more]

PMID: 28428565

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Project description:Involvement of the bacterial thiopurine methyltransferase (bTPMT) in natural selenium methylation by freshwater was investigated. A freshwater environment that had no known selenium contamination but exhibited reproducible emission of dimethyl selenide (DMSe) or dimethyl diselenide (DMDSe) when it was supplemented with an organic form of selenium [(methyl)selenocysteine] or an inorganic form of selenium (sodium selenite) was used. The distribution of the bTPMT gene (tpm) in the microflora was studied. Freshwater bacteria growing on 10 micro M sodium selenite and 10 micro M sodium selenate were isolated, and 4.5 and 10% of the strains, respectively, were shown by colony blot hybridization to hybridize with a Pseudomonas syringae tpm DNA probe. Ribotyping showed that these strains are closely related. The complete rrs sequence of one of the strains, designated Hsa.28, was obtained and analyzed. Its closest phyletic neighbor was found to be the Pseudomonas anguilliseptica rrs sequence. The Hsa.28 strain grown with sodium selenite or (methyl)selenocysteine produced significant amounts of DMSe and DMDSe. The Hsa.28 tpm gene was isolated by genomic DNA library screening and sequencing. BLASTP comparisons of the deduced Hsa.28 bTPMT sequence with P. syringae, Pseudomonas aeruginosa, Vibrio cholerae, rat, and human thiopurine methyltransferase sequences revealed that the levels of similarity were 52 to 71%. PCR-generated Escherichia coli subclones containing the Hsa.28 tpm open reading frame were constructed. E. coli cells harboring the constructs and grown with sodium selenite or (methyl)selenocysteine produced significant levels of DMSe and DMDSe, confirming that the gene plays a role in selenium methylation. The effect of strain Hsa.28 population levels on freshwater DMSe and DMDSe emission was investigated. An increase in the size of the Hsa.28 population was found to enhance significantly the emission of methyl selenides by freshwater samples supplemented with sodium selenite or (methyl)selenocysteine. These data suggest that bTPMT can play a role in natural freshwater selenium methylation processes.

Dataset Information

Small methyltransferase RlmH assembles a composite active site to methylate a ribosomal pseudouridine.

Publications

Small methyltransferase RlmH assembles a composite active site to methylate a ribosomal pseudouridine.

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