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Crystal structure of Escherichia coli protein ybgI, a toroidal structure with a dinuclear metal site.

Ladner JE, Obmolova G, Teplyakov A, Howard AJ, Khil PP, Camerini-Otero RD, Gilliland GL - BMC Struct. Biol. (2003)

Bottom Line: The protein encoded by the gene ybgI was chosen as a target for a structural genomics project emphasizing the relation of protein structure to function.The toroidal structure is comparable to that of some proteins that are involved in DNA metabolism.The di-nuclear metal site could imply that the specific function of this protein is as a hydrolase-oxidase enzyme.

View Article: PubMed Central - HTML - PubMed

Affiliation: Center for Advanced Research in Biotechnology, University of Maryland Biotechnology Institute and the National Institute of Standards and Technology, 9600 Gudelsky Drive, Rockville, MD 20850, USA. jane.ladner@nist.gov

ABSTRACT

Background: The protein encoded by the gene ybgI was chosen as a target for a structural genomics project emphasizing the relation of protein structure to function.

Results: The structure of the ybgI protein is a toroid composed of six polypeptide chains forming a trimer of dimers. Each polypeptide chain binds two metal ions on the inside of the toroid.

Conclusion: The toroidal structure is comparable to that of some proteins that are involved in DNA metabolism. The di-nuclear metal site could imply that the specific function of this protein is as a hydrolase-oxidase enzyme.

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The putative active site with the metal ions shown in silver balls and four water molecules shown as cyan balls. This figure was prepared using MOLSCRIPT [31] and Raster3D [32,33].
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Figure 3: The putative active site with the metal ions shown in silver balls and four water molecules shown as cyan balls. This figure was prepared using MOLSCRIPT [31] and Raster3D [32,33].

Mentions: The most likely region for the active site is a group of conserved residues which includes four histidines (63, 64, 97, 215), two glutamic acids (194, 219), one aspartic acid (101), one asparagine (108), one cysteine (171), one tyrosine (22) and one tryptophan (68). There are also two metal ions 3.3 Å apart in the selenomethionine protein and 2.5 Å apart in the native protein bound by this cluster of residues. In the early refinement of the selenomethionine structure these were treated as 'water' molecules and the B-values became very low indicating that they must be something heavier than oxygen. The anomalous Fourier map of the selenomethionine data indicates that there is a significant anomalous signal at these positions, though much lower than selenium. The X-ray fluorescence identified the presence of Fe in the protein sample. One metal ion is coordinated by H64 Ne2, H215 Ne2 and E219 Oe1; the other metal ion is coordinated by D101 Od1 and Od2, E219 Oe2 and H63 Ne2. This grouping is set back into the inside wall of the toroid and includes residues from both domains. The metal ion sites of the dimer are at opposite ends of a cavity that extends across the dimer interface. This cavity is separated from the center of the toroid by the Y22 residues of the dimer chains. The Y22 residues narrow the access to the cavity from the center of the toroid to approximately 14 Å The distances between the metal ions of the dimer chains are 21.9 and 25.6 Å. The distances between 3-fold related metal ions are 45.0 and 42.5 Å. One of the six putative active sites is shown in Figure 3. In the native protein, the metal positions may be filled or partially filled by magnesium ions which are present in both the growth medium and in the crystallization solution. An anomalous fourier using the native data does not reveal any anomalous signal at these positions and negative results using X-ray fluorescence eliminate the presence of Fe, Zn, Cu, Ni, and Co. In this structure, only 11 sites were included. The electron density at these positions tends to be somewhat smeared. The appearance of the electron density and the refinement of the B factors were used as guides to include or exclude ion sites. The protein structure around the sites is quite good. The presence of iron in the selenomethionine protein sample may indicate the adventitious uptake of iron during preparation since the procedure includes the addition of iron sulfate as a component in the growth medium [14]. The intrinsic metal ions for this protein are not known. The inclusion of histidine, glutamic acid, and aspartic acid in the putative active site with a bridging glutamic acid between the ions is in keeping with cocatalytic sites in a number of proteins where the metal ions are Zn or Zn and Fe, Mn, or Mg [15]. The constancy of the protein structure around these sites supports the view that these are catalytic rather than structural sites.


Crystal structure of Escherichia coli protein ybgI, a toroidal structure with a dinuclear metal site.

Ladner JE, Obmolova G, Teplyakov A, Howard AJ, Khil PP, Camerini-Otero RD, Gilliland GL - BMC Struct. Biol. (2003)

The putative active site with the metal ions shown in silver balls and four water molecules shown as cyan balls. This figure was prepared using MOLSCRIPT [31] and Raster3D [32,33].
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC239858&req=5

Figure 3: The putative active site with the metal ions shown in silver balls and four water molecules shown as cyan balls. This figure was prepared using MOLSCRIPT [31] and Raster3D [32,33].
Mentions: The most likely region for the active site is a group of conserved residues which includes four histidines (63, 64, 97, 215), two glutamic acids (194, 219), one aspartic acid (101), one asparagine (108), one cysteine (171), one tyrosine (22) and one tryptophan (68). There are also two metal ions 3.3 Å apart in the selenomethionine protein and 2.5 Å apart in the native protein bound by this cluster of residues. In the early refinement of the selenomethionine structure these were treated as 'water' molecules and the B-values became very low indicating that they must be something heavier than oxygen. The anomalous Fourier map of the selenomethionine data indicates that there is a significant anomalous signal at these positions, though much lower than selenium. The X-ray fluorescence identified the presence of Fe in the protein sample. One metal ion is coordinated by H64 Ne2, H215 Ne2 and E219 Oe1; the other metal ion is coordinated by D101 Od1 and Od2, E219 Oe2 and H63 Ne2. This grouping is set back into the inside wall of the toroid and includes residues from both domains. The metal ion sites of the dimer are at opposite ends of a cavity that extends across the dimer interface. This cavity is separated from the center of the toroid by the Y22 residues of the dimer chains. The Y22 residues narrow the access to the cavity from the center of the toroid to approximately 14 Å The distances between the metal ions of the dimer chains are 21.9 and 25.6 Å. The distances between 3-fold related metal ions are 45.0 and 42.5 Å. One of the six putative active sites is shown in Figure 3. In the native protein, the metal positions may be filled or partially filled by magnesium ions which are present in both the growth medium and in the crystallization solution. An anomalous fourier using the native data does not reveal any anomalous signal at these positions and negative results using X-ray fluorescence eliminate the presence of Fe, Zn, Cu, Ni, and Co. In this structure, only 11 sites were included. The electron density at these positions tends to be somewhat smeared. The appearance of the electron density and the refinement of the B factors were used as guides to include or exclude ion sites. The protein structure around the sites is quite good. The presence of iron in the selenomethionine protein sample may indicate the adventitious uptake of iron during preparation since the procedure includes the addition of iron sulfate as a component in the growth medium [14]. The intrinsic metal ions for this protein are not known. The inclusion of histidine, glutamic acid, and aspartic acid in the putative active site with a bridging glutamic acid between the ions is in keeping with cocatalytic sites in a number of proteins where the metal ions are Zn or Zn and Fe, Mn, or Mg [15]. The constancy of the protein structure around these sites supports the view that these are catalytic rather than structural sites.

Bottom Line: The protein encoded by the gene ybgI was chosen as a target for a structural genomics project emphasizing the relation of protein structure to function.The toroidal structure is comparable to that of some proteins that are involved in DNA metabolism.The di-nuclear metal site could imply that the specific function of this protein is as a hydrolase-oxidase enzyme.

View Article: PubMed Central - HTML - PubMed

Affiliation: Center for Advanced Research in Biotechnology, University of Maryland Biotechnology Institute and the National Institute of Standards and Technology, 9600 Gudelsky Drive, Rockville, MD 20850, USA. jane.ladner@nist.gov

ABSTRACT

Background: The protein encoded by the gene ybgI was chosen as a target for a structural genomics project emphasizing the relation of protein structure to function.

Results: The structure of the ybgI protein is a toroid composed of six polypeptide chains forming a trimer of dimers. Each polypeptide chain binds two metal ions on the inside of the toroid.

Conclusion: The toroidal structure is comparable to that of some proteins that are involved in DNA metabolism. The di-nuclear metal site could imply that the specific function of this protein is as a hydrolase-oxidase enzyme.

Show MeSH