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RNA:(guanine-N2) methyltransferases RsmC/RsmD and their homologs revisited--bioinformatic analysis and prediction of the active site based on the uncharacterized Mj0882 protein structure.

Bujnicki JM, Rychlewski L - BMC Bioinformatics (2002)

Bottom Line: Based on the results of sequence analysis and structure prediction, the residues involved in cofactor binding, target recognition and catalysis were identified, and the mechanism of the guanine-N2 methyltransfer reaction was proposed.Using the known Mj0882 structure, a comprehensive analysis of sequence-structure-function relationships in the family of genuine and putative m2G MTases was performed.The results provide novel insight into the mechanism of m2G methylation and will serve as a platform for experimental analysis of numerous uncharacterized N-MTases.

View Article: PubMed Central - HTML - PubMed

Affiliation: Bioinformatics Laboratory, International Institute of Cell and Molecular Biology, ul, ks, Trojdena 4, 02-109 Warsaw, Poland. iamb@wp.pl

ABSTRACT

Background: Escherichia coli guanine-N2 (m2G) methyltransferases (MTases) RsmC and RsmD modify nucleosides G1207 and G966 of 16S rRNA. They possess a common MTase domain in the C-terminus and a variable region in the N-terminus. Their C-terminal domain is related to the YbiN family of hypothetical MTases, but nothing is known about the structure or function of the N-terminal domain.

Results: Using a combination of sequence database searches and fold recognition methods it has been demonstrated that the N-termini of RsmC and RsmD are related to each other and that they represent a "degenerated" version of the C-terminal MTase domain. Novel members of the YbiN family from Archaea and Eukaryota were also indentified. It is inferred that YbiN and both domains of RsmC and RsmD are closely related to a family of putative MTases from Gram-positive bacteria and Archaea, typified by the Mj0882 protein from M. jannaschii (1dus in PDB). Based on the results of sequence analysis and structure prediction, the residues involved in cofactor binding, target recognition and catalysis were identified, and the mechanism of the guanine-N2 methyltransfer reaction was proposed.

Conclusions: Using the known Mj0882 structure, a comprehensive analysis of sequence-structure-function relationships in the family of genuine and putative m2G MTases was performed. The results provide novel insight into the mechanism of m2G methylation and will serve as a platform for experimental analysis of numerous uncharacterized N-MTases.

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Comparison of the conformations of the "catalytic loop" (motif IV) of N-MTases in stereoview. DNA:m6A MTases: M.TaqI (1g38 in PDB) [29] in yellow, M.RsrI (leg2) [55] in red, M.DpnM (2dpm) [56] in green; RNA:m6A MTase ErmC' (1qao) [24] in blue; DNA:m4C MTase M.PvuII (1boo) [57] in cyan; Mj0882 (1dus) in magenta. The ligand for each protein is shown: AdoMet for M.DpnM and ErmC', S-adenosylhomocysteine for M.PvuII, 5'-methylthioadenosine for M.RsrI, and 5'-[2-(amino)ethylthio]-5'-deoxyadenosine (cofactor analog) and the target deoxyadenosine for M.TaqI. In case of M.RsrI, ErmC' and M.PvuII, whose structures were solved in the absence of the target, the Tyr or Phe sidechain can be easily rotated to the M.TaqI-like orientation to make a face-to-face π-stacking interaction with the target base.
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Figure 2: Comparison of the conformations of the "catalytic loop" (motif IV) of N-MTases in stereoview. DNA:m6A MTases: M.TaqI (1g38 in PDB) [29] in yellow, M.RsrI (leg2) [55] in red, M.DpnM (2dpm) [56] in green; RNA:m6A MTase ErmC' (1qao) [24] in blue; DNA:m4C MTase M.PvuII (1boo) [57] in cyan; Mj0882 (1dus) in magenta. The ligand for each protein is shown: AdoMet for M.DpnM and ErmC', S-adenosylhomocysteine for M.PvuII, 5'-methylthioadenosine for M.RsrI, and 5'-[2-(amino)ethylthio]-5'-deoxyadenosine (cofactor analog) and the target deoxyadenosine for M.TaqI. In case of M.RsrI, ErmC' and M.PvuII, whose structures were solved in the absence of the target, the Tyr or Phe sidechain can be easily rotated to the M.TaqI-like orientation to make a face-to-face π-stacking interaction with the target base.

Mentions: The issue of divergent or convergent evolution notwithstanding, the "catalytic loop", corresponding to motif IV, assumes a strikingly similar conformation in all N-MTase structures solved to date (Figure 2). This suggests that the interactions between the target amino group and the side chain of the first residue and the carbonyl oxygen of the second residue of the tetrapeptide are highly similar. The same conformation is retained in the Mj0882 structure, consistent with the prediction that this protein belongs to the N-MTase family. Nevertheless, in the superimposed structures there is a substantial variation in the conformation of the cofactor AdoMet or its analogs, and in their relative orientation with respect to the catalytic site (Figure 2). This variation was noted previously in the course of crystallographic analyses, and was attributed to subtle structural differences between cofactor-binding pockets of various MTases that impose distinct conformation of the same ligand [12,24] and to the fact that different ligands (AdoMet, AdoHcy, and their analogs) make different interactions and do not necessarily retain the common conformation when bound to the same MTase [24,25].


RNA:(guanine-N2) methyltransferases RsmC/RsmD and their homologs revisited--bioinformatic analysis and prediction of the active site based on the uncharacterized Mj0882 protein structure.

Bujnicki JM, Rychlewski L - BMC Bioinformatics (2002)

Comparison of the conformations of the "catalytic loop" (motif IV) of N-MTases in stereoview. DNA:m6A MTases: M.TaqI (1g38 in PDB) [29] in yellow, M.RsrI (leg2) [55] in red, M.DpnM (2dpm) [56] in green; RNA:m6A MTase ErmC' (1qao) [24] in blue; DNA:m4C MTase M.PvuII (1boo) [57] in cyan; Mj0882 (1dus) in magenta. The ligand for each protein is shown: AdoMet for M.DpnM and ErmC', S-adenosylhomocysteine for M.PvuII, 5'-methylthioadenosine for M.RsrI, and 5'-[2-(amino)ethylthio]-5'-deoxyadenosine (cofactor analog) and the target deoxyadenosine for M.TaqI. In case of M.RsrI, ErmC' and M.PvuII, whose structures were solved in the absence of the target, the Tyr or Phe sidechain can be easily rotated to the M.TaqI-like orientation to make a face-to-face π-stacking interaction with the target base.
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Related In: Results  -  Collection

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Figure 2: Comparison of the conformations of the "catalytic loop" (motif IV) of N-MTases in stereoview. DNA:m6A MTases: M.TaqI (1g38 in PDB) [29] in yellow, M.RsrI (leg2) [55] in red, M.DpnM (2dpm) [56] in green; RNA:m6A MTase ErmC' (1qao) [24] in blue; DNA:m4C MTase M.PvuII (1boo) [57] in cyan; Mj0882 (1dus) in magenta. The ligand for each protein is shown: AdoMet for M.DpnM and ErmC', S-adenosylhomocysteine for M.PvuII, 5'-methylthioadenosine for M.RsrI, and 5'-[2-(amino)ethylthio]-5'-deoxyadenosine (cofactor analog) and the target deoxyadenosine for M.TaqI. In case of M.RsrI, ErmC' and M.PvuII, whose structures were solved in the absence of the target, the Tyr or Phe sidechain can be easily rotated to the M.TaqI-like orientation to make a face-to-face π-stacking interaction with the target base.
Mentions: The issue of divergent or convergent evolution notwithstanding, the "catalytic loop", corresponding to motif IV, assumes a strikingly similar conformation in all N-MTase structures solved to date (Figure 2). This suggests that the interactions between the target amino group and the side chain of the first residue and the carbonyl oxygen of the second residue of the tetrapeptide are highly similar. The same conformation is retained in the Mj0882 structure, consistent with the prediction that this protein belongs to the N-MTase family. Nevertheless, in the superimposed structures there is a substantial variation in the conformation of the cofactor AdoMet or its analogs, and in their relative orientation with respect to the catalytic site (Figure 2). This variation was noted previously in the course of crystallographic analyses, and was attributed to subtle structural differences between cofactor-binding pockets of various MTases that impose distinct conformation of the same ligand [12,24] and to the fact that different ligands (AdoMet, AdoHcy, and their analogs) make different interactions and do not necessarily retain the common conformation when bound to the same MTase [24,25].

Bottom Line: Based on the results of sequence analysis and structure prediction, the residues involved in cofactor binding, target recognition and catalysis were identified, and the mechanism of the guanine-N2 methyltransfer reaction was proposed.Using the known Mj0882 structure, a comprehensive analysis of sequence-structure-function relationships in the family of genuine and putative m2G MTases was performed.The results provide novel insight into the mechanism of m2G methylation and will serve as a platform for experimental analysis of numerous uncharacterized N-MTases.

View Article: PubMed Central - HTML - PubMed

Affiliation: Bioinformatics Laboratory, International Institute of Cell and Molecular Biology, ul, ks, Trojdena 4, 02-109 Warsaw, Poland. iamb@wp.pl

ABSTRACT

Background: Escherichia coli guanine-N2 (m2G) methyltransferases (MTases) RsmC and RsmD modify nucleosides G1207 and G966 of 16S rRNA. They possess a common MTase domain in the C-terminus and a variable region in the N-terminus. Their C-terminal domain is related to the YbiN family of hypothetical MTases, but nothing is known about the structure or function of the N-terminal domain.

Results: Using a combination of sequence database searches and fold recognition methods it has been demonstrated that the N-termini of RsmC and RsmD are related to each other and that they represent a "degenerated" version of the C-terminal MTase domain. Novel members of the YbiN family from Archaea and Eukaryota were also indentified. It is inferred that YbiN and both domains of RsmC and RsmD are closely related to a family of putative MTases from Gram-positive bacteria and Archaea, typified by the Mj0882 protein from M. jannaschii (1dus in PDB). Based on the results of sequence analysis and structure prediction, the residues involved in cofactor binding, target recognition and catalysis were identified, and the mechanism of the guanine-N2 methyltransfer reaction was proposed.

Conclusions: Using the known Mj0882 structure, a comprehensive analysis of sequence-structure-function relationships in the family of genuine and putative m2G MTases was performed. The results provide novel insight into the mechanism of m2G methylation and will serve as a platform for experimental analysis of numerous uncharacterized N-MTases.

Show MeSH
Related in: MedlinePlus