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ABSTRACT: The three-dimensional structure of ceruloplasmin (CP) with unoccupied labile metal-binding sites and the structure of CP containing
Ni2+ in the labile sites were solved for the first time at 2.6 and 2.95 Å resolution, respectively. Crystallization was performed
with the use of storage-stable CP, which was prepared in the presence of proteinase inhibitors and purified from (pre)proteinases.
Ceruloplasmin with Ni2+ crystallized in the orthorhombic space group, which had been earlier unknown for CP. Ceruloplasmin with the unoccupied labile
sites crystallized in the trigonal crystal form. The differences in intermolecular contacts observed in the trigonal and orthorhombic
crystal structures of CP are considered. The conformational changes attendant upon Ni2+ binding are described. It was suggested that the labile sites are multifunctional and can both bind metal ions potentially
toxic to organisms and be involved in electron transfer from substrates to the active site.
Crystallography Reports 07/2008; 53(4):655-662. · 0.47 Impact Factor
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ABSTRACT: Here, we describe high-resolution X-ray structures of Escherichia coli inorganic pyrophosphatase (E-PPase) complexed with the substrate, magnesium, or manganese pyrophosphate. The structures correspond to steps in the catalytic synthesis of enzyme-bound pyrophosphate (PP(i)) in the presence of fluoride as an inhibitor of hydrolysis. The catalytic reaction intermediates were trapped applying a new method that we developed for initiating hydrolytic activity in the E-PPase crystal. X-ray structures were obtained for three consecutive states of the enzyme in the course of hydrolysis. Comparative analysis of these structures showed that the Mn2+-supported hydrolysis of the phosphoanhydride bond is followed by a fast release of the leaving phosphate from the P1 site. The electrophilic phosphate P2 is trapped in the "down" conformation. Its movement into the "up" position most likely represents the rate-limiting step of Mn2+-supported hydrolysis. We further determined the crystal structure of the Arg43Gln mutant variant of E-PPase complexed with one phosphate and four Mn ions.
Journal of Molecular Biology 04/2007; 366(4):1305-17. · 4.00 Impact Factor
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ABSTRACT: The effect of relative humidity in a crystallization box on the rate of establishment of supersaturation conditions during
protein crystallization by diffusion of solvent vapors is estimated. A modified crystallization box is designed, which provides
the formation of a stable air flow with a specified relative humidity and its measurement directly in the closed space between
a drop and a reservoir. The range of relative humidities necessary to obtain the supersaturation conditions in a drop with
a protein crystallization solution is determined.
Crystallography Reports 04/2006; 51(3):519-524. · 0.47 Impact Factor
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ABSTRACT: The crystal structures of myoglobin in the deoxy- and carbon monoxide-ligated states at a resolution of 1.15 angstroms show that carbon monoxide binding at ambient temperatures requires concerted motions of the heme, the iron, and helices E and F for relief of steric inhibition. These steps constitute the main mechanism by which heme proteins lower the affinity of the heme group for the toxic ligand carbon monoxide.
Science 05/1999; 284(5413):473-6. · 31.20 Impact Factor
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J. Mol. Biol. 396(2010),785-799.
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[show abstract]
[hide abstract]
ABSTRACT: Here, we describe high-resolution X-ray structures of Escherichia coli inorganic pyrophosphatase (E-PPase) complexed with the substrate, magnesium, or manganese pyrophosphate. The structures correspond to steps in the catalytic synthesis of enzyme-bound pyrophosphate (PPi) in the presence of fluoride as an inhibitor of hydrolysis. The catalytic reaction intermediates were trapped applying a new method that we developed for initiating hydrolytic activity in the E-PPase crystal. X-ray structures were obtained for three consecutive states of the enzyme in the course of hydrolysis. Comparative analysis of these structures showed that the Mn2+-supported hydrolysis of the phosphoanhydride bond is followed by a fast release of the leaving phosphate from the P1 site. The electrophilic phosphate P2 is trapped in the “down” conformation. Its movement into the “up” position most likely represents the rate-limiting step of Mn2+-supported hydrolysis. We further determined the crystal structure of the Arg43Gln mutant variant of E-PPase complexed with one phosphate and four Mn ions.
Journal of Molecular Biology.
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Crystallogr. Rep. 51(2006),519-524.
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J. Mol. Biol. 364(2006),411-423.
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Biophysics, v.47, 795-805 (2002).
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J. Mol. Biol. 384(2008),489-502.
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Acta Crystallogr. F 63(2007),795-797.
