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Crystal structure of putative CbiT from Methanocaldococcus jannaschii: an intermediate enzyme activity in cobalamin (vitamin B12) biosynthesis.

Padmanabhan B, Yokoyama S, Bessho Y - BMC Struct. Biol. (2013)

Bottom Line: The overall tertiary structure and the tetrameric arrangements are highly homologous to those found in MT0146/CbiT from Methanobacterium thermoautotrophicum.The conservation of functional residues in the binding site for the co-factor, AdoMet, and in the putative precorrin-7 binding pocket suggested that MJ0391 may also possess CbiT activity.The putative function of MJ0391 is discussed, based on structural homology.

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Affiliation: Department of Biophysics, National Institute of Mental Health and Neuro Sciences (NIMHANS), Bangalore 560029, India. paddy@nimhans.kar.nic.in

ABSTRACT

Background: In the anaerobic pathway of cobalamin (vitamin B12) synthesis, the CbiT enzyme plays two roles, as a cobalt-precorrin-7 C15-methyltransferase and a C12-decarboxylase, to produce the intermediate, cobalt-precorrin 8.

Results: The primary structure of the hypothetical protein MJ0391, from Methanocaldococcus jannaschii, suggested that MJ0391 is a putative CbiT. Here, we report the crystal structure of MJ0391, solved by the MAD procedure and refined to final R-factor and R-free values of 19.8 & 27.3%, respectively, at 2.3 Å resolution. The asymmetric unit contains two NCS molecules, and the intact tetramer generated by crystallographic symmetry may be functionally important. The overall tertiary structure and the tetrameric arrangements are highly homologous to those found in MT0146/CbiT from Methanobacterium thermoautotrophicum.

Conclusions: The conservation of functional residues in the binding site for the co-factor, AdoMet, and in the putative precorrin-7 binding pocket suggested that MJ0391 may also possess CbiT activity. The putative function of MJ0391 is discussed, based on structural homology.

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Comparison with CbiT homologous proteins. (a) Sequence alignment of MJ0391 from Methanocaldococcus jannaschii, MT0146/CbiT from Methanobacterium thermoautotrophicum, precorrin-6Y C5, C15 – methyltransferase from Geobacter metallireducens GS-15 (Q39YF0), and the putative precorrin-6Y C5, C15 – methyltransferase from Corynebacterium diphtheriae (Q6NHA4). The secondary structural features from MJ0391 are indicated above the alignments. The colors reflect the similarity (red boxes and white characters for conserved residues; red characters for similarity in a group; blue frames for similarity across groups). The sequence was aligned and rendered by Clustal W [25] and ESPript [26], respectively. (b) Superposition of MJ0391 (yellow) on the precorrin-6Y C5, C15 – methyltransferase structures from G. metallireducens (PDB ID: 3E05, blue) and CobLc from R. capsulatus (PDB ID: 3NJR, cyan).
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Figure 3: Comparison with CbiT homologous proteins. (a) Sequence alignment of MJ0391 from Methanocaldococcus jannaschii, MT0146/CbiT from Methanobacterium thermoautotrophicum, precorrin-6Y C5, C15 – methyltransferase from Geobacter metallireducens GS-15 (Q39YF0), and the putative precorrin-6Y C5, C15 – methyltransferase from Corynebacterium diphtheriae (Q6NHA4). The secondary structural features from MJ0391 are indicated above the alignments. The colors reflect the similarity (red boxes and white characters for conserved residues; red characters for similarity in a group; blue frames for similarity across groups). The sequence was aligned and rendered by Clustal W [25] and ESPript [26], respectively. (b) Superposition of MJ0391 (yellow) on the precorrin-6Y C5, C15 – methyltransferase structures from G. metallireducens (PDB ID: 3E05, blue) and CobLc from R. capsulatus (PDB ID: 3NJR, cyan).

Mentions: A DALI [23] search revealed that although MJ0391 shares less sequence similarity with MT0146/CbiT from Methanobacterium thermoautotrophicum (33% identity; Z-score: 27.3), the precorrin-6Y C5, C15 – methyltransferase from Geobacter metallireducens GS-15 (Q39YF0; 32% identity; Z-score: 26.0), and the putative precorrin-6Y C5, C15 – methyltransferase from Corynebacterium diphtheriae (Q6NHA4; 17% identity; Z-score: 17.9) (Figure 3a), the overall tertiary structures and the quaternary structure arrangements are essentially conserved between them. The superimposition of the MJ0391 structure onto the structures of MT0146/CbiT (PDB ID: 1L3I), Q39YF0 (PDB ID: 3E05; 211-405aa), and Q6NHA4 (PDB ID: 3HM2; 236-410aa) yielded r.m.s.d. values of 1.6, 1.7, and 1.9 Å, respectively, for all Cα atoms (Figure 3b). Recently, the crystal structure of the C-terminal region of CobL (CobLc) from Rhodobacter capsulatus was reported [24]. The enzyme CobL, which uses precorrin-6B as a substrate, promotes the decarboxylation of the acetic acid side chain at C12, and the methylation at the C5 & C15 meso positions, to produce precorrin-8X. The superimposition of MJ0391 onto the CobLc structure (PDB ID: 3NJR) yielded Z-score and r.m.s.d. values of 25.6 and 1.8 Å, respectively, for 173 Cα atoms (Figure 3b).


Crystal structure of putative CbiT from Methanocaldococcus jannaschii: an intermediate enzyme activity in cobalamin (vitamin B12) biosynthesis.

Padmanabhan B, Yokoyama S, Bessho Y - BMC Struct. Biol. (2013)

Comparison with CbiT homologous proteins. (a) Sequence alignment of MJ0391 from Methanocaldococcus jannaschii, MT0146/CbiT from Methanobacterium thermoautotrophicum, precorrin-6Y C5, C15 – methyltransferase from Geobacter metallireducens GS-15 (Q39YF0), and the putative precorrin-6Y C5, C15 – methyltransferase from Corynebacterium diphtheriae (Q6NHA4). The secondary structural features from MJ0391 are indicated above the alignments. The colors reflect the similarity (red boxes and white characters for conserved residues; red characters for similarity in a group; blue frames for similarity across groups). The sequence was aligned and rendered by Clustal W [25] and ESPript [26], respectively. (b) Superposition of MJ0391 (yellow) on the precorrin-6Y C5, C15 – methyltransferase structures from G. metallireducens (PDB ID: 3E05, blue) and CobLc from R. capsulatus (PDB ID: 3NJR, cyan).
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Figure 3: Comparison with CbiT homologous proteins. (a) Sequence alignment of MJ0391 from Methanocaldococcus jannaschii, MT0146/CbiT from Methanobacterium thermoautotrophicum, precorrin-6Y C5, C15 – methyltransferase from Geobacter metallireducens GS-15 (Q39YF0), and the putative precorrin-6Y C5, C15 – methyltransferase from Corynebacterium diphtheriae (Q6NHA4). The secondary structural features from MJ0391 are indicated above the alignments. The colors reflect the similarity (red boxes and white characters for conserved residues; red characters for similarity in a group; blue frames for similarity across groups). The sequence was aligned and rendered by Clustal W [25] and ESPript [26], respectively. (b) Superposition of MJ0391 (yellow) on the precorrin-6Y C5, C15 – methyltransferase structures from G. metallireducens (PDB ID: 3E05, blue) and CobLc from R. capsulatus (PDB ID: 3NJR, cyan).
Mentions: A DALI [23] search revealed that although MJ0391 shares less sequence similarity with MT0146/CbiT from Methanobacterium thermoautotrophicum (33% identity; Z-score: 27.3), the precorrin-6Y C5, C15 – methyltransferase from Geobacter metallireducens GS-15 (Q39YF0; 32% identity; Z-score: 26.0), and the putative precorrin-6Y C5, C15 – methyltransferase from Corynebacterium diphtheriae (Q6NHA4; 17% identity; Z-score: 17.9) (Figure 3a), the overall tertiary structures and the quaternary structure arrangements are essentially conserved between them. The superimposition of the MJ0391 structure onto the structures of MT0146/CbiT (PDB ID: 1L3I), Q39YF0 (PDB ID: 3E05; 211-405aa), and Q6NHA4 (PDB ID: 3HM2; 236-410aa) yielded r.m.s.d. values of 1.6, 1.7, and 1.9 Å, respectively, for all Cα atoms (Figure 3b). Recently, the crystal structure of the C-terminal region of CobL (CobLc) from Rhodobacter capsulatus was reported [24]. The enzyme CobL, which uses precorrin-6B as a substrate, promotes the decarboxylation of the acetic acid side chain at C12, and the methylation at the C5 & C15 meso positions, to produce precorrin-8X. The superimposition of MJ0391 onto the CobLc structure (PDB ID: 3NJR) yielded Z-score and r.m.s.d. values of 25.6 and 1.8 Å, respectively, for 173 Cα atoms (Figure 3b).

Bottom Line: The overall tertiary structure and the tetrameric arrangements are highly homologous to those found in MT0146/CbiT from Methanobacterium thermoautotrophicum.The conservation of functional residues in the binding site for the co-factor, AdoMet, and in the putative precorrin-7 binding pocket suggested that MJ0391 may also possess CbiT activity.The putative function of MJ0391 is discussed, based on structural homology.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Biophysics, National Institute of Mental Health and Neuro Sciences (NIMHANS), Bangalore 560029, India. paddy@nimhans.kar.nic.in

ABSTRACT

Background: In the anaerobic pathway of cobalamin (vitamin B12) synthesis, the CbiT enzyme plays two roles, as a cobalt-precorrin-7 C15-methyltransferase and a C12-decarboxylase, to produce the intermediate, cobalt-precorrin 8.

Results: The primary structure of the hypothetical protein MJ0391, from Methanocaldococcus jannaschii, suggested that MJ0391 is a putative CbiT. Here, we report the crystal structure of MJ0391, solved by the MAD procedure and refined to final R-factor and R-free values of 19.8 & 27.3%, respectively, at 2.3 Å resolution. The asymmetric unit contains two NCS molecules, and the intact tetramer generated by crystallographic symmetry may be functionally important. The overall tertiary structure and the tetrameric arrangements are highly homologous to those found in MT0146/CbiT from Methanobacterium thermoautotrophicum.

Conclusions: The conservation of functional residues in the binding site for the co-factor, AdoMet, and in the putative precorrin-7 binding pocket suggested that MJ0391 may also possess CbiT activity. The putative function of MJ0391 is discussed, based on structural homology.

Show MeSH
Related in: MedlinePlus