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Nitrogenase MoFe protein from Clostridium pasteurianum at 1.08 Å resolution: comparison with the Azotobacter vinelandii MoFe protein.

Zhang LM, Morrison CN, Kaiser JT, Rees DC - Acta Crystallogr. D Biol. Crystallogr. (2015)

Bottom Line: The surrounding environment is also highly conserved, suggesting that this structural arrangement is crucial for nitrogen reduction.The P clusters are likewise similar, although the surrounding protein and solvent environment is less conserved relative to that of the FeMo cofactor.This makes it plausible that this loop is repositioned to open up the Fe protein docking surface for complex formation.

View Article: PubMed Central - HTML - PubMed

Affiliation: Division of Chemistry and Chemical Engineering, California Institute of Technology, Pasadena, CA 91125, USA.

ABSTRACT
The X-ray crystal structure of the nitrogenase MoFe protein from Clostridium pasteurianum (Cp1) has been determined at 1.08 Å resolution by multiwavelength anomalous diffraction phasing. Cp1 and the ortholog from Azotobacter vinelandii (Av1) represent two distinct families of nitrogenases, differing primarily by a long insertion in the α-subunit and a deletion in the β-subunit of Cp1 relative to Av1. Comparison of these two MoFe protein structures at atomic resolution reveals conserved structural arrangements that are significant to the function of nitrogenase. The FeMo cofactors defining the active sites of the MoFe protein are essentially identical between the two proteins. The surrounding environment is also highly conserved, suggesting that this structural arrangement is crucial for nitrogen reduction. The P clusters are likewise similar, although the surrounding protein and solvent environment is less conserved relative to that of the FeMo cofactor. The P cluster and FeMo cofactor in Av1 and Cp1 are connected through a conserved water tunnel surrounded by similar secondary-structure elements. The long α-subunit insertion loop occludes the presumed Fe protein docking surface on Cp1 with few contacts to the remainder of the protein. This makes it plausible that this loop is repositioned to open up the Fe protein docking surface for complex formation.

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Ribbon representation of the Cp1 tetramer viewed down the molecular (noncrystallographic) twofold symmetry axis. The α-subunits are shaded in pale cyan and teal, except for the long insertion loops (residues α376–α429), which are highlighted in blue. The β-subunits are colored split pea and pale green. The FeMo cofactors and the P clusters are depicted as sticks, with C atoms shown in light gray, N atoms in blue, O atoms in red, S atoms in yellow, Fe atoms in orange and Mo atoms in cyan.
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fig1: Ribbon representation of the Cp1 tetramer viewed down the molecular (noncrystallographic) twofold symmetry axis. The α-subunits are shaded in pale cyan and teal, except for the long insertion loops (residues α376–α429), which are highlighted in blue. The β-subunits are colored split pea and pale green. The FeMo cofactors and the P clusters are depicted as sticks, with C atoms shown in light gray, N atoms in blue, O atoms in red, S atoms in yellow, Fe atoms in orange and Mo atoms in cyan.

Mentions: The 1.08 Å resolution X-ray crystal structure of Cp1 was determined by experimental phasing using MAD data (see §2 for details). The crystal form of Cp1 is similar to the previously reported cesium-derivative Cp1 structure in space group P21, with unit-cell parameters a = 72.7, b = 170.6, c = 87.5 Å, β = 91.6° (Kim et al., 1993 ▶). The data-processing and refinement statistics are summarized in Tables 1 ▶ and 2 ▶, respectively. An overview of the Cp1 tetramer structure is shown in Fig. 1 ▶. A total of 1953 residues (out of 1982 residues) and 2484 water molecules were included in the model for structural refinement. The first two residues in the N-terminus and the last 11–12 residues in the C-terminus of the α-subunit were not included in the model owing to weak electron density. The overall coordinate error in the model is estimated to be 0.022 Å from the diffraction-component precision index (Cruickshank, 1999 ▶). A single nonproline cis-peptide bond, first reported in Kp1 between Trpα251 and Serα252, is also present in Cp1 and Av1 (Leuα240–Thrα241 in Cp1 and Trpα253–Serα254 in Av1; Mayer et al., 1999 ▶; Spatzal et al., 2011 ▶). This pair of residues is located in a β-sheet of the second domain in the α-subunit (Kim & Rees, 1992 ▶), about 9 Å away from the FeMo cofactor. A few registry errors were corrected from the previously reported 3.0 Å resolution Cp1 structure, among which the longest (residues 412–420) is within the α-subunit insertion sequence characteristic of the group II MoFe proteins (Supplementary Fig. S1; Kim et al., 1993 ▶).


Nitrogenase MoFe protein from Clostridium pasteurianum at 1.08 Å resolution: comparison with the Azotobacter vinelandii MoFe protein.

Zhang LM, Morrison CN, Kaiser JT, Rees DC - Acta Crystallogr. D Biol. Crystallogr. (2015)

Ribbon representation of the Cp1 tetramer viewed down the molecular (noncrystallographic) twofold symmetry axis. The α-subunits are shaded in pale cyan and teal, except for the long insertion loops (residues α376–α429), which are highlighted in blue. The β-subunits are colored split pea and pale green. The FeMo cofactors and the P clusters are depicted as sticks, with C atoms shown in light gray, N atoms in blue, O atoms in red, S atoms in yellow, Fe atoms in orange and Mo atoms in cyan.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4321486&req=5

fig1: Ribbon representation of the Cp1 tetramer viewed down the molecular (noncrystallographic) twofold symmetry axis. The α-subunits are shaded in pale cyan and teal, except for the long insertion loops (residues α376–α429), which are highlighted in blue. The β-subunits are colored split pea and pale green. The FeMo cofactors and the P clusters are depicted as sticks, with C atoms shown in light gray, N atoms in blue, O atoms in red, S atoms in yellow, Fe atoms in orange and Mo atoms in cyan.
Mentions: The 1.08 Å resolution X-ray crystal structure of Cp1 was determined by experimental phasing using MAD data (see §2 for details). The crystal form of Cp1 is similar to the previously reported cesium-derivative Cp1 structure in space group P21, with unit-cell parameters a = 72.7, b = 170.6, c = 87.5 Å, β = 91.6° (Kim et al., 1993 ▶). The data-processing and refinement statistics are summarized in Tables 1 ▶ and 2 ▶, respectively. An overview of the Cp1 tetramer structure is shown in Fig. 1 ▶. A total of 1953 residues (out of 1982 residues) and 2484 water molecules were included in the model for structural refinement. The first two residues in the N-terminus and the last 11–12 residues in the C-terminus of the α-subunit were not included in the model owing to weak electron density. The overall coordinate error in the model is estimated to be 0.022 Å from the diffraction-component precision index (Cruickshank, 1999 ▶). A single nonproline cis-peptide bond, first reported in Kp1 between Trpα251 and Serα252, is also present in Cp1 and Av1 (Leuα240–Thrα241 in Cp1 and Trpα253–Serα254 in Av1; Mayer et al., 1999 ▶; Spatzal et al., 2011 ▶). This pair of residues is located in a β-sheet of the second domain in the α-subunit (Kim & Rees, 1992 ▶), about 9 Å away from the FeMo cofactor. A few registry errors were corrected from the previously reported 3.0 Å resolution Cp1 structure, among which the longest (residues 412–420) is within the α-subunit insertion sequence characteristic of the group II MoFe proteins (Supplementary Fig. S1; Kim et al., 1993 ▶).

Bottom Line: The surrounding environment is also highly conserved, suggesting that this structural arrangement is crucial for nitrogen reduction.The P clusters are likewise similar, although the surrounding protein and solvent environment is less conserved relative to that of the FeMo cofactor.This makes it plausible that this loop is repositioned to open up the Fe protein docking surface for complex formation.

View Article: PubMed Central - HTML - PubMed

Affiliation: Division of Chemistry and Chemical Engineering, California Institute of Technology, Pasadena, CA 91125, USA.

ABSTRACT
The X-ray crystal structure of the nitrogenase MoFe protein from Clostridium pasteurianum (Cp1) has been determined at 1.08 Å resolution by multiwavelength anomalous diffraction phasing. Cp1 and the ortholog from Azotobacter vinelandii (Av1) represent two distinct families of nitrogenases, differing primarily by a long insertion in the α-subunit and a deletion in the β-subunit of Cp1 relative to Av1. Comparison of these two MoFe protein structures at atomic resolution reveals conserved structural arrangements that are significant to the function of nitrogenase. The FeMo cofactors defining the active sites of the MoFe protein are essentially identical between the two proteins. The surrounding environment is also highly conserved, suggesting that this structural arrangement is crucial for nitrogen reduction. The P clusters are likewise similar, although the surrounding protein and solvent environment is less conserved relative to that of the FeMo cofactor. The P cluster and FeMo cofactor in Av1 and Cp1 are connected through a conserved water tunnel surrounded by similar secondary-structure elements. The long α-subunit insertion loop occludes the presumed Fe protein docking surface on Cp1 with few contacts to the remainder of the protein. This makes it plausible that this loop is repositioned to open up the Fe protein docking surface for complex formation.

Show MeSH
Related in: MedlinePlus