[DYNAMICS AND MECHANISMS OF INTERACTION OF HETERO-HEXAMERIC TIP49a/b COMPLEXES WITH DS-DNA].

Evolutionary conserved TIP49a and TIP49b ATPases belong to the AAA+ superfamily of DNA-dependent ATPases that are involved in many cellular processes such as chromatin remodeling, regulation of transcription and cell division during mitosis, the maintenance of genome stability, snoRNP biogenesis, and participate in the formation of active form of telomerase. These proteins are involved in the complex networks of protein-protein interactions and, in spite of high structural similarity, in some cases, can perform opposite functions. Despite of the variety of their different activities, the exact mechanisms of action of TIP49a and TIP49b are still poorly understood. In this paper, by means of molecular docking approaches we first modeled the structures of hetero-hexameric TIP49 complexes with short ds-DNA fragments (20 base pairs with different GC content) within the central channel of hexameric ring. Using molecular dynamics simulations in the periodic water box (MD) we investigated conformational dynamics and mechanisms of DNA unwinding activity of these proteins. We shown that: a) the interaction between the positively charged protein loops and DNA within the central channel of protein ring leads to the partial unwinding of the DNA helix; b) DNA unwinding occurs only in the region within the protein ring, while the terminal parts of DNA outside the protein complex remain in its initial b-form conformation; c) the presence of ATP in the active sites of protein complex affects both the dynamics and the structure of DNA, leading to the breakage of some complementary bonds in AT-rich DNA sequences.

X-ray beam induced current method at the laboratory x-ray source.

The x-ray beam induced current method (XBIC) is realized on the laboratory x-ray source using the polycapillary x-ray optics. It is shown that rather good images of grain boundaries in Si can be obtained by this method. The parameters of x-ray beam are estimated by the simulation of Schottky diode image. A good correlation between the experimental and calculated grain boundary XBIC contrast is obtained. The possibilities of laboratory source based XBIC method are estimated.

X-ray nanointerferometer based on si refractive bilenses.

We report a novel type of x-ray interferometer employing a bilens system consisting of two parallel compound refractive lenses, each of which creates a diffraction limited beam under coherent illumination. By closely overlapping such coherent beams, an interference field with a fringe spacing ranging from tens of nanometers to tens of micrometers is produced. In an experiment performed with 12 keV x rays, submicron fringes were observed by scanning and moiré imaging of the test grid. The far field interference pattern was used to characterize the x-ray coherence. Our technique opens up new opportunities for studying natural and man-made nanoscale materials.

Thermodynamics and structure of actinide(IV) complexes with nitrilotriacetic acid.

Nitrilotriacetic acid, commonly known as NTA (N(CH(2)CO(2)H)(3)), can be considered a representative of the polyaminocarboxylic family. The results presented in this paper describe the thermodynamical complexation and structural investigation of An(IV) complexes with NTA in aqueous solution. In the first part, the stability constants of the An(IV) complexes (An = Pu, Np, U, and Th) have been determined by spectrophotometry. In the second part, the coordination spheres of the actinide cation in these complexes have been described using extended X-ray absorption fine structure spectroscopy and compared to the solid-state structure of (Hpy)(2)[U(NTA)(2)] x (H(2)O). These data are further compared to quantum chemical calculations, and their evolution across the actinide series is discussed. In particular, an interpretation of the role of the nitrogen atom in the coordination mode is proposed. These results are considered to be model behavior of polyaminocarboxylic ligands such as diethylenetriamine pentaacetic acid, which is nowadays the best candidate for a chelating agent in the framework of actinide decorporation for the human body.

Two-step hard X-ray focusing combining Fresnel zone plate and single-bounce ellipsoidal capillary.

A two-step focusing set-up combining a Fresnel zone plate with an ellipsoidal capillary is presented. It is shown that, in addition to the anticipated gain in flux, the employment of the prefocusing micro-optic makes optimal use of the elliptical shape of the capillary by almost eliminating aberrations. A small cross section of the prefocused beam allows a tiny fraction of the capillary surface to be selected, thus reducing the influence of slope errors. An X-ray beam with a 15 keV energy was focused down to a spot size as small as 250 nm, demonstrating the best value that has been achieved up to now for single-bounce capillaries. The use of an ellipsoidal capillary as a micromirror under off-axis illumination by microfocusing optics may open up new opportunities in nanofocusing developments.

Submicrometer hard X-ray focusing using a single-bounce ellipsoidal capillary combined with a Fresnel zone plate.

A single-bounce capillary with an ellipsoidal shape has been used for two-step focusing in combination with a Fresnel zone plate (FZP). The FZP serves as a first microfocusing element and produces a demagnified micrometer image of the source, before the elliptical capillary makes a last final compression of the beam. With 15 keV X-rays from the European Synchrotron Radiation Facility BM5 bending magnet, the two-step demagnification system produced a focus of about 250 nm with a gain of more than 1000. The use of an ellipsoidal capillary as a micro-mirror under off-axis illumination using micro-prefocusing optics might open up new opportunities in nanofocusing developments.

Single-molecule study of RuvAB-mediated Holliday-junction migration.

Branch migration of Holliday junctions is an important step of genetic recombination and DNA repair. In Escherichia coli, this process is driven by the RuvAB complex acting as a molecular motor. Using magnetic tweezers, we studied the RuvAB-directed migration of individual Holliday junctions formed between two approximately 6-kb DNA molecules of identical sequence, and we measured the migration rate at 37 degrees C and 1 mM ATP. We directly demonstrate that RuvAB is a highly processive DNA motor protein that is able to drive continuous and unidirectional branch migration of Holliday junctions at a well defined average speed over several kilobases through homologous sequences. We observed directional inversions of the migration at the DNA molecule boundaries leading to forth-and-back migration of the branch point and allowing us to measure the migration rate in the presence of negative or positive loads. The average migration rate at zero load was found to be approximately 43 bp/sec. Furthermore, the load dependence of the migration rate is small, within the force range of -3.4 pN (hindering force) to +3.4 pN (assisting force).

Distinct prevalence of the CYP19 Delta3(TTTA)(7) allele in premenopausal versus postmenopausal breast cancer patients, but not in control individuals.

The CYP19 gene encodes the enzyme aromatase, which plays a key role in the conversion of androgens to oestrogens. A polymorphism in CYP19 in intron 4 (TTTA)n has been reported to be associated with breast cancer (BC) risk, although conflicting evidence has also been published. Here, we employ a non-traditional, highly demonstrative design of a molecular epidemiological study, where the comparison of BC cases and healthy middle-aged female donors was supplemented by an analysis of groups with extreme characteristics of either BC risk (bilateral breast cancer (biBC) patients) or cancer tolerance (tumour-free elderly women aged >or=75 years). None of the (TTTA)n polymorphic variants was significantly overrepresented among the affected women compared with any of the control groups. However, a 3-bp deletion/insertion CYP19 polymorphism, which is located in the same intron approximately 50 bp upstream to the (TTTA)n repeat, was evidently associated with the menopausal status in both the BC and biBC cohorts. In particular, the Delta3(TTTA)(7) allele occurred significantly more frequently in premenopausal than in postmenopausal BC patients (65/172 (38%) versus 67/310 (22%); P=0.0001; Odds Ratio (OR)=2.20 (95% Confidence Interval (CI) 1.46-3.32)), while the perimenopausal cases demonstrated an intermediate value (9/34 (26%)). In the biBC cohort, women who developed both tumours during their premenopausal period had a significantly higher prevalence of the Delta3(TTTA)(7) allele than patients with a postmenopausal onset of bilateral disease (16/46 (35%) versus 8/50 (16%); P=0.035; OR=2.80 (1.08-7.23)); those biBC patients, whose tumours were diagnosed before and after the cessation of menses, displayed an intermediate occurrence of the Delta3(TTTA)(7) allele (7/32 (22%)). Similar tendencies in the Delta3(TTTA)(7) allele distribution in BC and biBC patients suggest that its association with the menopausal status of the patients is truly non-random and thus this observation deserves further detailed investigation.

Pyridinium tetrakis(nitrato-kappa2O,O')(2,2':6',2"-terpyridine-kappa3N)cerate(III) pyridine solvate and bis(methanol-kappaO)tris(nitrato-kappa2O,O')(2,2':6',2"-terpyridine-kappa3N)cerium(III).

The title complexes, (C(5)H(6)N)[Ce(NO(3))(4)(C(15)H(11)N(3))].C(5)H(5)N or (Hpy)[Ce(NO(3))(4)(terpy)].py, (I) (py is pyridine, C(5)H(5)N, and terpy is terpyridine, C(15)H(11)N(3)), and [Ce(NO(3))(3)(C(15)H(11)N(3))(CH(4)O)(2)] or [Ce(NO(3))(3)(terpy)(OHCH(3))(2)], (II), are 11-coordinate. The coordination polyhedron of the Ce atom in (I) is irregular, while that in (II) can be described as an icosahedron with two vertices replaced by one.

Tissue transglutaminase expression in breast carcinomas.

Tissue transglutaminase (tTG) is known to participate in multiple cellular processes, including apoptosis, cellular adhesiveness etc. Alterations of tTG expression could contribute to the development of several categories of diseases, including AIDS, cancer etc. The aim of the study was to test the pattern and relevance of tTG expression in a subset of breast carcinomas. RT-PCR has detected tTG-specific RNA message in 11 out of 25 (44%) breast cancer samples. tTG message was detected in 6/8 (75%) breast carcinomas with high apoptotic index, but only in 5/17 (29%) with the low one (p = 0.03). Immunohistochemical analysis revealed that only 15% of breast carcinomas displayed tTG protein in tumor cells, while the staining of the stromal components occurred in approximately one-half of the tumours tested. Surprisingly, there was no significant association between tTG RNA expression and protein positivity. Moreover, there was no evident relationships between tTG immunostaining and apoptotic index or clinical parameters of breast neoplasms. There are at least 2 alternative explanations for the poor concordance between RNA and protein data. It is likely that the sensitivity of immunohistochemistry is not sufficient to detect functionally relevant tTG enzyme in all breast cancer sections. Otherwise, tTG RNA expression does not always lead to accumulation of its product in the tumor cells, but reflects the transcriptional activation of other pro-apoptotic genes due to common triggering mechanisms.

CYP19 gene polymorphism in endometrial cancer patients.

PURPOSE: Initiation/promotion of endometrial cancer is known to be associated with estrogenic influence. Therefore, it is possible that some allelic polymorphisms of the genes involved in steroidogenesis or steroid metabolism contribute to endometrial cancer susceptibility. METHODS: Here, we compared CYP19 (aromatase) gene polymorphism in 85 endometrial cancer patients and in 110 non-affected women. RESULTS: The genotypes containing the longest alleles (A6 and A7) of CYP19 were found to be over-represented in patients as compared to controls. In addition, these genotypes demonstrated a tendency to be associated with increased concentrations of estradiol and testosterone in postmenopausal patients. CONCLUSIONS: Thus, CYP19 polymorphism might be one of the genetic risk factors for endometrial cancer development.

Involvement of the TIP60 histone acetylase complex in DNA repair and apoptosis.

It is well known that histone acetylases are important chromatin modifiers and that they play a central role in chromatin transcription. Here, we present evidence for novel roles of histone acetylases. The TIP60 histone acetylase purifies as a multimeric protein complex. Besides histone acetylase activity on chromatin, the TIP60 complex possesses ATPase, DNA helicase, and structural DNA binding activities. Ectopic expression of mutated TIP60 lacking histone acetylase activity results in cells with defective double-strand DNA break repair. Importantly, the resulting cells lose their apoptotic competence, suggesting a defect in the cells' ability to signal the existence of DNA damage to the apoptotic machinery. These results indicate that the histone acetylase TIP60-containing complex plays a role in DNA repair and apoptosis.

CYP17 polymorphism in the groups of distinct breast cancer susceptibility: comparison of patients with the bilateral disease vs. monolateral breast cancer patients vs. middle-aged female controls vs. elderly tumor-free women.

The CYP17 gene encodes an enzyme involved in several critical steps of steroidogenesis. The promoter region of the CYP17 displays a single-nucleotide polymorphism, which is suspected to modulate the expression of the gene and thus may contribute in the interindividual variations of hormonal background. In agreement with this functional hypothesis, the MspA1+ allele (designated as A2) of the CYP17 was shown to render an increased risk of breast cancer (BC). However, the latter observation was disputed by a series of negative reports. Here, we re-evaluated the role of CYP17 MspA1 polymorphism in the BC susceptibility, using a non-traditional design of a case-control study. In addition to randomly selected 183 BC patients and 107 female middle-aged donors, we examined the groups with apparently extreme characteristics of either BC risk or BC resistance, namely the 57 bilateral breast cancer (biBC) patients and 75 elderly (>/=75 years old) tumor-free women. Neither BC nor biBC patients showed increased prevalence of 'unfavorable' A2 allele as compared with the non-affected cohorts. Moreover, the A2 variant was not significantly associated with the tumor size, nodal involvement and menopausal status in the patients either with the monolateral or bilateral disease. Thus, our data argue against the earlier reported role of the CYP17 in BC predisposition and progression. In addition, usual distribution of the CYP17 alleles in the elderly group indicates a neutral effect of this polymorphism on the longevity in females.

Evidence for microsatellite instability in bilateral breast carcinomas.

The molecular pathogenesis of various categories of breast cancer (BC) has been well described, but surprisingly few reports have appeared on analysis of somatic mutations in bilateral BC. We have performed a polymerase chain reaction (PCR)-driven investigation of chromosomal regions showing common loss of heterozygosity (LOH) in 23 cases (46 tumors) from patients diagnosed with bilateral BC. LOH was observed in 15/46 (33%) informative tumors for chromosome 1p, 5/32 (16%) for 5q, 12/44 (27%) for 11q, 15/40 (38%) for 13q and 4/24 (17%) for 17p. These values are within the range of interlaboratory variations reported for unilateral BC. There was no strong evidence for concordance of LOH within the same patient for any of the chromosomal loci tested. Atypical for breast carcinomas, 7/46 (15%) tumors accumulated a high frequency (ranging from 11 to 29%) of shortened dinucleotide CA repeats, implying microsatellite instability (MI). Further analysis with the highly informative BAT-26 marker allowed for the classification of two of these tumors as having a replication error positive (RER(+)/MSI-H) phenotype, whereas the remaining five carcinomas harbored so-called borderline MI. Thus an involvement of both RER(+) and borderline MI appears to be a distinct feature of bilateral breast carcinomas compared to unilateral lesions.

A cruciform structural transition provides a molecular switch for chromosome structure and dynamics.

The interaction between specific sites along a DNA molecule is often crucial for the regulation of genetic processes. However, mechanisms regulating the interaction of specific sites are unknown. We have used atomic force microscopy to demonstrate that the structural transition between cruciform conformations can act as a molecular switch to facilitate or prevent communication between distant regions in DNA. Cruciform structures exist in vivo and they are critically involved in the initiation of replication and the regulation of gene expression in different organisms. Therefore, structural transitions of the cruciform may play a key role in these processes.

L-myc polymorphism in cancer patients, healthy blood donors and elderly, tumor-free individuals in Russia.

L-myc polymorphism was investigated in 95 breast cancer (BC), 63 colorectal cancer (CC) and 58 lung cancer (LC) patients, as well as in 122 healthy, middle-aged blood donors (HBDs) and 184 elderly, tumor-free individuals. The occurrence of the S allele in the BC cohort (57%) was significantly higher than that in middle-aged, healthy females (41%) and elderly, non-affected women (47%), implying involvement of the L-myc genotype in BC susceptibility (age-adjusted OR = 1.74, 95% CI 1.11-2.73, p = 0.016). L-myc allele distribution in CC and LC was similar to that in controls. Contrary to earlier reports, L:S allele frequencies ratio in elderly blood donors (EBDs) did not significantly differ from that in HBDs (0.49:0. 51 and 0.54:0.46, respectively). However, the S allele had a tendency to be over-represented among elderly compared with middle-aged smokers (55% vs. 44%; OR = 1.57, 95% CI 0.98-2.50, p = 0. 059), which implies that it may be linked with tolerance to smoking effects.

Crystallographic, X-ray absorption, and IR studies of solid- and solution-state structures of tris(nitrato) N,N,N',N'-tetraethylmalonamide complexes of lanthanides. Comparison with the Americium complex.

To fine-tune the design of optimized donor ligands for nuclear waste actinide selective extraction, both electronic and molecular structures of the actinide complexes that are formed must be investigated. In particular, to achieve the selective complexation of transplutonium 3+ ions versus lanthanide 3+ ions is one of the major challenges, given the chemical similarities between these two f-element families. In this work, the structure of solvent-phase M(NO3)3(TEMA)2 complexes (Ln = Nd, Eu, Ho, Yb, Lu, Am; TEMA = N,N,N',N'-tetraethylmalonamide) was investigated by liquid-phase spectroscopic methods among which extended X-ray absorption fine structure played a major role. In addition, the crystal structures of the species Nd(NO3)3(TEMA)2 and Yb(NO3)3(TEMA)2 have been determined by X-ray diffraction. Nd(NO3)3(C11N2O2H22)2 crystallizes in the monoclinic system (P2(1) space group; a = 11.2627(4) A, b = 20.5992(8) A, c = 22.2126(8) A; alpha = gamma = 90 degrees, beta = 102.572(1) degrees; Z = 6), and Yb(NO3)3(C11N2O2H22)2 crystallizes in the orthorhombic system (P2(1)2(1)2(1) space group; a = 9.3542(1) A, b = 18.1148(2) A, c = 19.7675(2) A; alpha = beta = gamma = 90 degrees; Z = 4). In the solvent phase, the metal polyhedron was found to be similar to that of the solid-state complex Nd(NO3)3(TEMA)2 for M = Nd to Ho. For M = Yb and Lu, a significant elongation of one nitrate oxygen bond was observed. Comparison with measurements on the Am(NO3)3(TEMA)2 complex in ethanol has shown the similarities between the Nd3+ and Am3+ coordination spheres.

Migration of a Holliday junction through a nucleosome directed by the E. coli RuvAB motor protein.

Chromatin plays a critical role in regulating access to DNA by proteins that direct recombination and repair. The E. coli RuvAB protein complex promotes branch migration of the Holliday junction recombination intermediate. The ability of RuvAB to negotiate passage of the junction through nucleosomal DNA is examined. The model system involves the formation of a Holliday junction positioned upstream of a nucleosome. Unassisted, the junction is blocked by a histone octamer. In the presence of RuvAB and ATP, rapid branch migration through the nucleosome is observed. It results in disruption of the histone-DNA interactions leading to the removal of the octamer from the junction intermediate. These results suggest that eukaryotic DNA motor proteins analogous to RuvAB could function during recombination to promote branch migration through chromatin.