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Biomedical subjects

A N Popov

Publications and source records attributed to A N Popov.

At least 19 recordsLinked to original sources
PubMed2006

Reversible inhibition of Escherichia coli inorganic pyrophosphatase by fluoride: trapped catalytic intermediates in cryo-crystallographic studies.

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.

Binding Sites↗
PubMed2006

Crystallization and preliminary X-ray analysis of cytochrome c nitrite reductase from Thioalkalivibrio nitratireducens.

A novel cytochrome c nitrite reductase (TvNiR) was isolated from the haloalkalophilic bacterium Thioalkalivibrio nitratireducens. The enzyme catalyses nitrite and hydroxylamine reduction, with ammonia as the only product of both reactions. It consists of 525 amino-acid residues and contains eight haems c. TvNiR crystals were grown by the hanging-drop vapour-diffusion technique. The crystals display cubic symmetry and belong to space group P2(1)3, with unit-cell parameter a = 194 A. A native data set was obtained to 1.5 A resolution. The structure was solved by the SAD technique using the data collected at the Fe absorption peak wavelength.

Crystallization↗
PubMed2002

[Blood level of lipid peroxidation products as an indicator of resistance to infectious inflammatory complications of transurethral electroresection of the prostate].

Study of relationship between the content of lipid peroxidation (LPO) products in the blood of patients with benign prostatic hyperplasia (BPH) and resistance to infectious inflammatory complications (IIC) of transurethral electroresection of the prostate showed that decreased content of circulating lipoperoxides promoted the development of postoperative IIC. Before the operation blood levels of LPO products were increased in BPH patients who did not develop IIC postoperation in comparison with normal controls. Three intravenous injections of ascorbic acid in a single dose of 1000 mg after transurethral electroresection of the prostate led to an increase in the blood level of LPO products and promoted a decrease in the incidence of postoperative IIC.

Antioxidants↗
PubMed2001

The structures of Escherichia coli inorganic pyrophosphatase complexed with Ca(2+) or CaPP(i) at atomic resolution and their mechanistic implications.

Two structures of Escherichia coli soluble inorganic pyrophosphatase (EPPase) complexed with calcium pyrophosphate (CaPP(i)-EPPase) and with Ca(2+) (Ca(2+)-EPPase) have been solved at 1.2 and 1.1 A resolution, respectively. In the presence of Mg(2+), this enzyme cleaves pyrophosphate (PP(i)) into two molecules of orthophosphate (P(i)). This work has enabled us to locate PP(i) in the active site of the inorganic pyrophosphatases family in the presence of Ca(2+), which is an inhibitor of EPPase.Upon PP(i) binding, two Ca(2+) at M1 and M2 subsites move closer together and one of the liganded water molecules becomes bridging. The mutual location of PP(i) and the bridging water molecule in the presence of inhibitor cation is catalytically incompetent. To make a favourable PP(i) attack by this water molecule, modelling of a possible hydrolysable conformation of PP(i) in the CaPP(i)-EPPase active site has been performed. The reasons for Ca(2+) being the strong PPase inhibitor and the role in catalysis of each of four metal ions are the mechanistic aspects discussed on the basis of the structures described.

Binding Sites↗
PubMed2001

Morphometric parameters of inflammatory infiltration in renal tissue and personality characteristics of patients at risk of pyelonephritis attack after percutaneous nephrolithotomy.

A correlation between personality traits and the type of inflammatory infiltration in the renal pelvicalyceal system was revealed in patients with nephrolithiasis. The study demonstrated the possibility of indirect evaluation of the inflammatory process in the pelvicalyceal system and preoperative prediction of acute pyelonephritis attack after percutaneous nephrolithotomy on the basis of patient's psychological status.

Humans↗
PubMed2000

Improving the X-ray resolution by reversible flash-cooling combined with concentration screening, as exemplified with PPase.

A significant improvement in the X-ray resolution of crystals of Escherichia coli inorganic pyrophosphatase at cryotemperature was obtained as a result of studying the relationship between the crystal order and cryosolution component concentrations. To perform the experiments, the ability to reverse the flash-cooling process and to return a crystal to ambient temperature was used. In each cycle, the crystal was transferred from a cold nitrogen-gas stream to a cryosolution with modified concentrations of the components. The crystal was then flash-cooled again and the diffraction quality checked. Such a technique allows the screening of a wide concentration range rather quickly without using a large number of crystals and allows the determination of optimal cryosolution component concentrations. The resolution limit for crystals of pyrophosphatase increased by almost 0.7 A, from 1.8 to 1.15 A.

Crystallography, X-Ray↗
PubMed1999

A steric mechanism for inhibition of CO binding to heme proteins.

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.

Animals↗
PubMed1996

The binding of carbon monoxide and nitric oxide to leghaemoglobin in comparison with other haemoglobins.

Haemoglobins have the ability to discriminate between oxygen and other diatomic molecules. To further understanding of this process the X-ray crystal structures of carbonmonoxy and nitrosyl-leghaemoglobin have been determined at 1.8 A resolution. The ligand geometry is discussed in detail and the controversial issue of bent versus linear carbon monoxide binding is addressed. The bond angle of 160 degrees for CO-leghaemoglobin is in conflict with recent spectroscopy results on myoglobin but is consistent with angles obtained for myoglobin X-ray crystal structures. In contrast to the numerous carbon monoxide studies, very little stereochemical information is available for the nitric oxide adduct of haemoglobin. This is provided by the X-ray structure of NO-leghaemoglobin, which conforms to expected geometry with an Fe-NO angle of 147 degrees and a lengthened iron-proximal histidine bond. Thus crystallographic evidence is given for the predicted weakening of this bond on the binding of nitric oxide.

Binding Sites↗
PubMed1996

[A method for extrafocal controllable wound closure and an adaptable repositioning device].

For quicker healing of vast long-standing wounds the authors propose a specially developed device-wound contractors accelerating processes of contraction. Experimental investigations have shown that processes of concentric contraction depend on the shape and are most typical of wounds having the appearance of extended rectangle. Promising clinical results were obtained in treatment of 17 patients with vast defects of tissues of different localization (trunk--10, femur--3, shoulder--2, head--2).

Adolescent↗
PubMed1996

[Experimental and clinical validation of auto- and allodermoplasty using transverse dermal micrografts in deep burns].

A new method of autoallodermoplasty is proposed which is as follows: a patient's skin band is cut out 40 times less than the total area of the wound surface. The wound in the donor area is sutured tightly. The skin flap is dissected to thin transverse rectangular pieces with a special microdermatome or with a razor and then they are laid with their lateral side onto the wound and covered with the allo-skin. Experimental and clinical observations have shown that at remote terms both the anatomical and functional restitution of the integument occurred. The method is indicated for huge dermal defects of the "functionally passive" areas in patients with restricted, not big resources of the donor sites, especially in patients with high operative risk.

Animals↗
PubMed1996

[Anaerobic gas gangrene after the amputation of an extremity].

During 30 years of practical work the authors observed 6 patients with anaerobic mephitic gangrene after amputation of the extremity for atherosclerotic (in 3 patients), diabetic (in 2 patients) and vast wounds of the foot. Among the most substantial noninfective causes of the development of anaerobic mephitic gangrene in the postoperative period there were continuous preoperative ischemia of the extremity against the background of a decreased level of haemoglobin, total protein, lymphocytes. An early operation and intensive complex specific and nonspecific therapy resulted in recovery of 3 patients.

Aged↗
PubMed1995

The structure of deoxy- and oxy-leghaemoglobin from lupin.

The leghaemoglobins have oxygen affinities 11 to 24 times higher than that of sperm whale myoglobin, due mainly to higher rates of association. To find out why, we have determined the structures of deoxy- and oxy-leghaemoglobin II of the lupin at 1.7 A resolution. Results confirm the general features found in previous X-ray analyses of this protein. The unique feature that has now emerged is the rotational freedom of the proximal histidine. In deoxy-leghaemoglobin the imidazole oscillates between two alternative orientations, eclipsing either the lines N1-N3 or N2-N4 of the porphyrin; in oxy-leghaemoglobin it is fixed in a staggered orientation. The iron atom moves from a position 0.30 A from the plane of the pyrrole nitrogen atoms in deoxy- to a position in the plane in oxy-leghaemoglobin while the Fe- bond distance remains constant at 2.02 A. The Fe-O-O angle is 152 degrees, as in human haemoglobin. The oxygen is hydrogen-bonded to the distal histidine at N epsilon 2-O1 and N epsilon 2-O2 distance of 2.95 A and 2.68 A, respectively. The porphyrin is ruffled equally in deoxy- and oxy-leghaemoglobins, due to rotations of the pyrrols about the N-Fe-N bonds, causing the methine bridges to deviate by up to 0.32 A from the mean porphyrin plane. The only feature capable of accounting for the high on-rate of the reaction with oxygen are the mobilities of the proximal histidine and distal histidine residues in deoxy-leghaemoglobin. The eclipsed positions of the proximal histidine in deoxy-leghaemoglobin maximize steric hindrance with the porphyrin nitrogen atoms and minimize pi-->p electron donation, while its staggered position in oxy-leghaemoglobin reverses both these effects. Together with the oscillation of the imidazole between the two orientations, these two factors may reduce the activation energy for the reaction of leghaemoglobin with oxygen. The distal histidine is in a fixed position in the haem pocket in the crystal, but must be swinging in and out of the pocket at a high rate in solution to allow the oxygen to enter.

Crystallography, X-Ray↗
PubMed1995

Atomic structure at 2.5 A resolution of uridine phosphorylase from E. coli as refined in the monoclinic crystal lattice.

Uridine phosphorylase from E. coli (Upase) has been crystallized using vapor diffusion technique in a new monoclinic crystal form. The structure was determined by the molecular replacement method at 2.5 A resolution. The coordinates of the trigonal crystal form were used as a starting model and the refinement by the program XPLOR led to the R-factor of 18.6%. The amino acid fold of the protein was found to be the same as that in the trigonal crystals. The positions of flexible regions were refined. The conclusion about the involvement in the active site is in good agreement with the results of the biochemical experiments.

Amino Acid Sequence↗
PubMed1994

X-ray crystallographic studies of recombinant inorganic pyrophosphatase from Escherichia coli.

An E. coli inorganic pyrophosphatase overproducer and a method for a large-scale production of the homogeneous enzyme are described. The inorganic pyrophosphatase was crystallized in the form containing one subunit of a homohexameric molecule per asymmetric unit: space group R32, a = 110.4 A, c = 76.8 A. The electron density map to 2.5 A resolution phased with Eu- and Hg-derivatives (figure of merit, = 0.51) was improved by the solvent flattening procedure ( = 0.77). The course of the polypeptide chain and the secondary structure elements, intersubunit contacts and positions of the active sites were characterized. Homology with S. cerevisiae inorganic pyrophosphatase structure was found.

Binding Sites↗
PubMed1989

[The submolecular structure of the Carnation Mottle virus].

The model of the tertiary structure of the CMtV subunit was suggested and the mode of its packaging into the capsid was determined on the basis of electron microscopy small-angle X-ray scattering and X-ray diffraction data. The i-ray diffraction data on the crystal of the native virion of Carnation Mottle Virus were obtained on the automatic diffractometer supplied with two-dimensional detector.

Plant Viruses↗