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The non-canonical hydroxylase structure of YfcM reveals a metal ion-coordination motif required for EF-P hydroxylation.

Kobayashi K, Katz A, Rajkovic A, Ishii R, Branson OE, Freitas MA, Ishitani R, Ibba M, Nureki O - Nucleic Acids Res. (2014)

Bottom Line: The structure of YfcM is similar to that of the ribonuclease YbeY, even though they do not share sequence homology.Our findings showed that the metal ion-coordinating motif of YfcM plays an essential role in the hydroxylation of the β-lysylated lysine residue of EF-P.Taken together, our results suggested the potential catalytic mechanism of hydroxylation by YfcM.

View Article: PubMed Central - PubMed

Affiliation: Department of Biological Sciences, Graduate School of Science, The University of Tokyo, 2-11-16 Yayoi, Bunkyo-ku, Tokyo 113-0032, Japan Global Research Cluster, RIKEN, 2-1, Hirosawa, Wako, Saitama, 351-0198, Japan.

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Comparison of the metal-coordination motifs of YbeY and YfcM. (A) Metal-coordination motif of YfcM. The Fo-Fc omit map contoured at 10σ is shown in green. Disordered loops are represented by dashed lines. Metal ions are depicted as spheres, labeled M. Electrostatic interactions are represented by dashed blue lines. (B) Superimposition of the structures of YfcM and YbeY. The proteins are color-coded as in Figure 2. Residues are depicted by ball-and-stick models.
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Figure 4: Comparison of the metal-coordination motifs of YbeY and YfcM. (A) Metal-coordination motif of YfcM. The Fo-Fc omit map contoured at 10σ is shown in green. Disordered loops are represented by dashed lines. Metal ions are depicted as spheres, labeled M. Electrostatic interactions are represented by dashed blue lines. (B) Superimposition of the structures of YfcM and YbeY. The proteins are color-coded as in Figure 2. Residues are depicted by ball-and-stick models.

Mentions: In the structure of YbeY, the three-histidine motif located on α5 and the loop between α5 and α6 coordinates a metal ion (Figure 3A). The α5 and α6 helices of YbeY structurally correspond to the α2 and α3 helices of YfcM, respectively (Figure 2A, B and C). Our structure of YfcM revealed that it also coordinates a metal ion with His59 and His63 on α2, and Glu98 on α3 (Figure 4A). The metal ion is probably derived from the E. coli lysate, but its identity is unclear. Interestingly, the structural superimposition of YbeY and YfcM revealed the nearly identical positions of the metal ion-coordinating residues of these two proteins, with His59, His63 and Glu98 of YfcM corresponding to His114, His118 and His124 of YbeY, respectively (Figure 4B). In a similar manner, Glu137 of YbeY interacts with His114 and helps coordinate a metal ion. Likewise, Glu137 corresponds to Asp105 of YfcM, which interacts with His59 (Figure 4B). The interaction between Asp 105 and His59 may help coordinate Glu98 to the metal ion by bringing α3 into closer proximity to α2. A sequence alignment of the metal binding motif revealed that His59, His63 and Glul98 are all highly conserved in YfcM (Supplementary Figure S2). Asp105 is also conserved as the acidic residue (aspartate or glutamate) (Supplementary Figure S2). Therefore, YfcM harbors a metal ion-coordinating motif consisting of two histidines and a glutamate, and their positions overlap with those of the metal ion-coordinating motif in YbeY.


The non-canonical hydroxylase structure of YfcM reveals a metal ion-coordination motif required for EF-P hydroxylation.

Kobayashi K, Katz A, Rajkovic A, Ishii R, Branson OE, Freitas MA, Ishitani R, Ibba M, Nureki O - Nucleic Acids Res. (2014)

Comparison of the metal-coordination motifs of YbeY and YfcM. (A) Metal-coordination motif of YfcM. The Fo-Fc omit map contoured at 10σ is shown in green. Disordered loops are represented by dashed lines. Metal ions are depicted as spheres, labeled M. Electrostatic interactions are represented by dashed blue lines. (B) Superimposition of the structures of YfcM and YbeY. The proteins are color-coded as in Figure 2. Residues are depicted by ball-and-stick models.
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Figure 4: Comparison of the metal-coordination motifs of YbeY and YfcM. (A) Metal-coordination motif of YfcM. The Fo-Fc omit map contoured at 10σ is shown in green. Disordered loops are represented by dashed lines. Metal ions are depicted as spheres, labeled M. Electrostatic interactions are represented by dashed blue lines. (B) Superimposition of the structures of YfcM and YbeY. The proteins are color-coded as in Figure 2. Residues are depicted by ball-and-stick models.
Mentions: In the structure of YbeY, the three-histidine motif located on α5 and the loop between α5 and α6 coordinates a metal ion (Figure 3A). The α5 and α6 helices of YbeY structurally correspond to the α2 and α3 helices of YfcM, respectively (Figure 2A, B and C). Our structure of YfcM revealed that it also coordinates a metal ion with His59 and His63 on α2, and Glu98 on α3 (Figure 4A). The metal ion is probably derived from the E. coli lysate, but its identity is unclear. Interestingly, the structural superimposition of YbeY and YfcM revealed the nearly identical positions of the metal ion-coordinating residues of these two proteins, with His59, His63 and Glu98 of YfcM corresponding to His114, His118 and His124 of YbeY, respectively (Figure 4B). In a similar manner, Glu137 of YbeY interacts with His114 and helps coordinate a metal ion. Likewise, Glu137 corresponds to Asp105 of YfcM, which interacts with His59 (Figure 4B). The interaction between Asp 105 and His59 may help coordinate Glu98 to the metal ion by bringing α3 into closer proximity to α2. A sequence alignment of the metal binding motif revealed that His59, His63 and Glul98 are all highly conserved in YfcM (Supplementary Figure S2). Asp105 is also conserved as the acidic residue (aspartate or glutamate) (Supplementary Figure S2). Therefore, YfcM harbors a metal ion-coordinating motif consisting of two histidines and a glutamate, and their positions overlap with those of the metal ion-coordinating motif in YbeY.

Bottom Line: The structure of YfcM is similar to that of the ribonuclease YbeY, even though they do not share sequence homology.Our findings showed that the metal ion-coordinating motif of YfcM plays an essential role in the hydroxylation of the β-lysylated lysine residue of EF-P.Taken together, our results suggested the potential catalytic mechanism of hydroxylation by YfcM.

View Article: PubMed Central - PubMed

Affiliation: Department of Biological Sciences, Graduate School of Science, The University of Tokyo, 2-11-16 Yayoi, Bunkyo-ku, Tokyo 113-0032, Japan Global Research Cluster, RIKEN, 2-1, Hirosawa, Wako, Saitama, 351-0198, Japan.

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Related in: MedlinePlus