[Combination of tumors of different histogenesis in pediatric neurosurgery]. / Sochetanie opukholei razlichnogo gistogeneza v detskoi neirokhirurgii.

Multiple gliomas are determined by synchronous two or more tumors located in different brain regions. It is important to distinguish multiple primary tumors and metastatic brain lesion. In the first case, tumor spread can`t be explained by dissemination along the cerebrospinal fluid pathways, commissural fibers or local metastases. Multiple primary tumors with different histological structures are called bidermal neoplasms. Surgery is preferred in these patients with severe symptoms. The purpose of surgery is maximum resection of tumor. Follow-up may be advisable for small tumors without clinical manifestations. Treatment of multiple gliomas includes surgery, radiotherapy and chemotherapy. Multiple tumor process in children is much more severe compared to a single neoplasia that requires neurological and neuroimaging control and determines treatment strategy. The authors report 3 children with multicentric gliomas, discuss the various aspects of diagnosis and treatment of multiple gliomas and formulate the recommendations for the treatment based on own clinical experience and literature data.

Substrate Profile of the Phosphotriesterase Homology Protein from Escherichia coli.

The phosphotriesterase homology protein (PHP) from Escherichia coli is a member of a family of proteins that is related to phosphotriestrase (PTE), a bacterial enzyme from cog1735 with unusual substrate specificity toward the hydrolysis of synthetic organic phosphates and phosphonates. PHP was cloned, purified to homogeneity, and functionally characterized. The three-dimensional structure of PHP was determined at a resolution of 1.84 Å with zinc and phosphate in the active site. The protein folds as a distorted (ß/α)8-barrel and possesses a binuclear metal center in the active site. The catalytic function and substrate profile of PHP were investigated using a structure-guided approach that combined bioinformatics, computational docking, organic synthesis, and steady-state enzyme kinetics. PHP was found to catalyze the hydrolysis of phosphorylated glyceryl acetates. The best substrate was 1,2-diacetyl glycerol-3-phosphate with a kcat/ Km of 4.9 × 103 M-1 s-1. The presence of a phosphate group in the substrate was essential for enzymatic hydrolysis by the enzyme. It was surprising, however, to find that PHP was unable to hydrolyze any of the lactones tested as potential substrates, unlike most of the other enzymes from cog1735.

Bioinformatics and molecular modeling in chemical enzymology. Active sites of hydrolases.

Comparison and multiple alignments of amino acid sequences of a representative number of related enzymes demonstrate the existence of certain positions of amino acid residues which are permanently reproducible in all members of the whole family. The use of the bioinformatic approach revealed conservative residues in each of the related enzymes and ranked amino acid conservatism for the overall enzymatic catalysis. Glycine and aspartic acid residues were shown to be the most essential for structure and catalytic activity of enzymes. Amino acid residues forming catalytic subsite of the active site of enzymes are always highly conservative. Analysis revealed that aspartic acid carboxyl group is the most frequently employed nucleophilic (in deprotonated form) and electrophilic (in protonated form) agent involved in activation of molecules by the mechanism of general base and acidic catalyses in the catalytic sites of enzymes. Glycine is a unique amino acid possessing the highest possibilities for rotation along C-C and C-N bonds of the polypeptide chain. The conservative fixation of the glycine residue in polypeptide chains of related enzymes provides a possibility for directed assembly of amino acid residues into the catalytic subsite structure. It is possible that the conservative glycines provide known conformational mobility of the protein and the active site. Methods of molecular modeling were used for analysis of structural substitutions of conservative and non-conservative glycines and their effects on geometry of catalytic site of typical hydrolases. The substitution of glycine(s) for alanine significantly altered the catalytic site structures.

Mapping the functional surface of domain 2 in the gelsolin superfamily.

The crystal structure of the F-actin binding domain 2 of severin, the gelsolin homologue from Dictyostelium discoideum, has been determined by multiple isomorphous replacement and refined to 1.75 A resolution. The structure reveals an alpha-helix-beta-sheet sandwich similar to the domains of gelsolin and villin, and contains two cation-binding sites, as observed in other domain 1 and domain 2 homologues. Comparison of the structures of several gelsolin family domains has identified residues that may mediate F-actin binding in gelsolin domain 2 homologues. To assess the involvement of these residues in F-actin binding, three mutants of human gelsolin domain 2 were assayed for F-actin binding activity and thermodynamic stability. Two of the mutants, RRV168AAA and RLK210AAA, demonstrated a lowered affinity for F-actin, indicating a role for those residues in filament binding. Using both structural and biochemical data, we have constructed a model of the gelsolin domain 1-domain 2-F-actin complex. This model highlights a number of interactions that may serve as positive and negative determinants of filament end- and side-binding.

Essential functions and actin-binding surfaces of yeast cofilin revealed by systematic mutagenesis.

Cofilin stimulates actin filament turnover in vivo. The phenotypes of twenty yeast cofilin mutants generated by systematic mutagenesis were determined. Ten grew as well as the wild type and showed no cytoskeleton defects, seven were recessive-lethal and three were conditional-lethal and caused severe actin organization defects. Biochemical characterization of interactions between nine mutant yeast cofilins and yeast actin provided evidence that F-actin binding and depolymerization are essential cofilin functions. Locating the mutated residues on the yeast cofilin molecular structure allowed several important conclusions to be drawn. First, residues required for actin monomer binding are proximal to each other. Secondly, additional residues are required for interactions with actin filaments; these residues might bind an adjacent subunit in the actin filament. Thirdly, despite striking structural similarity, cofilin interacts with actin in a different manner from gelsolin segment-1. Fourthly, a previously unrecognized cofilin function or interaction is suggested by identification of spatially proximal residues important for cofilin function in vivo, but not for actin interactions in vitro. Finally, mutation of the cofilin N-terminus suggests that its sequence is conserved because of its critical role in actin interactions, not because it is sometimes a target for protein kinases.

Structure determination of yeast cofilin.

Cofilin, a ubiquitous 15,000 M(r) protein, plays a central role in regulating cytoskeletal dynamics. Cofilin binds to actin monomers and filaments, and has a pH-dependent actin severing activity. The structure will allow for a detailed analysis of cofilin function.