[PCR analysis of the expression of chromosome 18 genes in human liver tissue: interindividual variability]. / PTsR-analiz ékspressii genov khromosomy 18 v tkani pecheni cheloveka: mezhindividual'naia variabel'nost'.

Using human chromosome 18 (Ch18) genes as an example, a PCR analysis of the interindividual variability of gene expression in liver tissue was performed. Although the quantitative profiles of the Ch18 transcriptome, expressed in the number of cDNA copies per single cell, showed a high degree of correlation between donors (Pearson correlation coefficients ranged from 0.963 to 0.966), the expression of the significant number of genes (from 13% to 19%, depending on the method of experimental data normalization) varied by more than 4-fold when comparing donors pairwise. At the same time, the proportion of differentially expressed genes increased with a decrease in the level of their expression. It is shown that the higher quantitative variability of low-abundance transcripts is mainly not technical, but biological. Bioinformatic analysis of the interindividual variability of the differential expression of chromosome 18 genes in human liver tissue did not reveal any statistically significant groups of genes related to certain biological processes that indicated a rather transient nature of the interindividual variability of their expression, probably reflecting the response of cells of an individual to specific external stimuli.

[Effects of the H6R and D7H Mutations on the Heparin-Dependent Modulation of Zinc-Induced Aggregation of Amyloid ß].

Zinc ions and glycosaminoglycans (GAGs) are found in amyloid deposits and are known to modulate the ß-amyloid peptide (Aß) aggregation, which is thought to be a key event in the pathogenesis of Alzheimer's disease (AD). Correlation spectroscopy was used to study how the H6R and D7H mutations of the metal-binding domain (MBD) of Aß42 affect the modulation of its zinc-induced aggregation by the model GAG heparin. The H6R mutation was shown to decrease and the D7H mutation to increase the Aß42 propensity to aggregate in the presence of zinc ions. In addition, H6R diminished and D7H enhanced the modulating effect of heparin. The difference in the heparin-dependent modulation was associated with coordination of zinc ions within the MBDs of the mutant peptides. The findings indicate that anion-binding sites formed by complexes of zinc ions with the Aß MBD play an essential role in the interaction of zinc-induced Aß aggregates with heparin.

[Evaluation of the diversity of random DNA-libraries by the shape of amplification curves for estimation of the efficiency of aptamer selection]. / Otsenka raznoobraziia kombinatornykh DNK-bibliotek na osnove analiza formy amplifikatsionnykh krivykh dlia monitoringa éffektivnosti selektsii aptamerov.

Using random (combinatorial) DNA-libraries with various degrees of diversity, it was shown that their amplification by polymerase chain reaction in real time resulted in appearance of a maximum on amplification curves. The relative decrease of fluorescence after passing the maximum was directly proportional to the logarithm of the number of oligonucleotide sequence variants in the random DNA-library provided that this number was within in the interval from 1 to 104 and remained practically unaltered when the number of variants was in the interval from 105 to 108. The obtained dependence was used in the course of SELEX to evaluate changes in the diversity of random DNA-libraries from round to round in selection of DNA-aptamers to the recombinant SMAD4 protein. As a result, oligonucleotides containing sequences able to form a site of SMAD4-DNA interactions known as SBE (SMAD-binding element) have been selected thus indicating that the SMAD4-SBE interaction dominates the aptamer selection.

[Zinc-induced interactions of the metal-binding domain of beta-amyloid with nucleic acids and glycosaminoglycans].

Zinc ions form complexes with ß-amyloid peptides and play an important role in Alzheimer's disease pathogenesis. It has been demonstrated by turbidimetry and correlation spectroscopy that synthetic peptide Aß16 representing the metal-binding domain of ß-amyloid is able to interact with nucleic acids, chondroitin polysulfate, and dextran sulfates in the presence of zinc ions. The amino acid D7H substitution enhanced the peptide binding to polyanions, whereas the H6R and H6A-H13A substitutions abolished this interaction. It is suggested that the metal-binding domain may serve as a zinc-dependent site of ß-amyloid interaction with biological polyanions including DNA, RNA, and glycosaminoglycans.

[Effect of mutations and modifications of amino acid residues on zinc-induced interaction of the metal-binding domain of ß-amyloid with DNA].

Interaction of intranuclear ß-amyloid with DNA is considered to be a plausible mechanism of Alzheimer's disease pathogenesis. The interaction of single- and double-stranded DNA with synthetic peptides was analyzed using surface plasmon resonance. The peptides represent the metal-binding domain of ß-amyloid (amino acids 1-16) and its variants with chemical modifications and point substitutions of amino acid residues which are associated with enhanced neurotoxicity of ß-amyloid in cell tests. It has been shown that the presence of zinc ions is necessary for the interaction of the peptides with DNA in solution. H6R substitution has remarkably reduced the ability of domain 1-16 to bind DNA. This is in accordance with the supposition that the coordination of a zinc ion by amino acid residues His6, Glu11, His13, and His14 of the ß-amyloid metal-binding domain results in the occurrence of an anion-binding site responsible for the interaction of the domain with DNA. Zinc-induced dimerization and oligomerization of domain 1-16 associated with phosphorylation of Ser8 and the presence of unblocked amino- and carboxy-terminal groups have resulted in a decrease of peptide concentrations required for detection of the peptide-DNA interaction. The presence of multiple anion-binding sites on the dimers and oligomers is responsible for the enhancement of the peptide-DNA interaction. A substitution of the negatively charged residue Asp7 for the neutral residue Asn in close proximity to the anion-binding site of the domain 1-16 of Aß facilitates the electrostatic interaction between this site and phosphates of a polynucleotide chain, which enhances zinc-induced binding to DNA.

[Physico-chemical methods for studing ß-amyloid aggregation].

Alzheimer's disease is the most prevalent neurodegenerative pathology. According to the amyloid cascade hypothesis, a key event of the Alzheimer's disease pathogenesis is a transition of the ß-amyloid peptide (Аß) from the monomeric form to the aggregated state. The mechanism of Ðß aggregation is intensively studied in vitro, by means of synthetic peptides and various physico-chemical methods allowing evaluation of size, molecular structure, and morphology of the formed aggregates. The paper reviews both the well-known and recently introduced physico-chemical methods for analysis of Ðß aggregation, including microscopу, optical and fluorescent methods, method of electron paramagnetic resonance, electrochemical and electrophoretic methods, gel-filtration, and mass spectrometric methods. Merits and drawbacks of the methods are discussed. The unique possibility to simultaneously observe Ðß monomers as well oligomers and large aggregates by means of atomic force microscopy or fluorescence correlation spectroscopy is emphasized. The high detection sensitivity of the latter method, monitoring the aggregation process in Ðß solutions at low peptide concentrations is underlined. Among mass spectrometric methods, the ion mobility mass spectrometry is marked out as a method enabling to obtain information about both the spectrum of Ðß oligomers and their structure. It is pointed out that the use of several methods giving the complementary data about Ðß aggregates is the best experimental approach to studying the process of b-amyloid peptide aggregation in vitro.