Mária Takács

Reconstruction of quaternary structure from X-ray scattering by equilibrium mixtures of biological macromolecules.

A recent renaissance in small-angle X-ray scattering (SAXS) made this technique a major tool for the low-resolution structural characterization of biological macromolecules in solution. The major limitation of existing methods for reconstructing 3D models from SAXS is imposed by the requirement of solute monodispersity. We present a novel approach that couples low-resolution 3D SAXS reconstruction with composition analysis of mixtures. The approach is applicable to polydisperse and difficult to purify systems, including weakly associated oligomers and transient complexes. Ab initio shape analysis is possible for symmetric homo-oligomers, whereas rigid body modeling is applied also to dissociating complexes when atomic structures of the individual subunits are available. In both approaches, the sample is considered as an equilibrium mixture of intact complexes/oligomers with their dissociation products or free subunits. The algorithms provide the 3D low-resolution model (for ab initio modeling, also the shape of the monomer) and the volume fractions of the bound and free state(s). The simultaneous fitting of multiple scattering data sets collected under different conditions allows one to restrain the modeling further. The possibilities of the approach are illustrated in simulated and experimental SAXS data from protein oligomers and multisubunit complexes including nucleoproteins. Using this approach, new structural insights are provided in the association behavior and conformations of estrogen-related receptors ERRα and ERRγ. The possibility of 3D modeling from the scattering by mixtures significantly widens the range of applicability of SAXS and opens novel avenues in the analysis of oligomeric mixtures and assembly/dissociation processes.

The Asymmetric Binding of PGC-1α to the ERRα and ERRγ Nuclear Receptor Homodimers Involves a Similar Recognition Mechanism

Background PGC-1α is a crucial regulator of cellular metabolism and energy homeostasis that functionally acts together with the estrogen-related receptors (ERRα and ERRγ) in the regulation of mitochondrial and metabolic gene networks. Dimerization of the ERRs is a pre-requisite for interactions with PGC-1α and other coactivators, eventually leading to transactivation. It was suggested recently (Devarakonda et al) that PGC-1α binds in a strikingly different manner to ERRγ ligand-binding domains (LBDs) compared to its mode of binding to ERRα and other nuclear receptors (NRs), where it interacts directly with the two ERRγ homodimer subunits. Methods/Principal Findings Here, we show that PGC-1α receptor interacting domain (RID) binds in an almost identical manner to ERRα and ERRγ homodimers. Microscale thermophoresis demonstrated that the interactions between PGC-1α RID and ERR LBDs involve a single receptor subunit through high-affinity, ERR-specific L3 and low-affinity L2 interactions. NMR studies further defined the limits of PGC-1α RID that interacts with ERRs. Consistent with these findings, the solution structures of PGC-1α/ERRα LBDs and PGC-1α/ERRγ LBDs complexes share an identical architecture with an asymmetric binding of PGC-1α to homodimeric ERR. Conclusions/Significance These studies provide the molecular determinants for the specificity of interactions between PGC-1α and the ERRs, whereby negative cooperativity prevails in the binding of the coactivators to these receptors. Our work indicates that allosteric regulation may be a general mechanism controlling the binding of the coactivators to homodimers.

Comparison of spinal curvature parameters as determined by the ZEBRIS spine examination method and the Cobb method in children with scoliosis

Background and purpose The most common and gold standard method to diagnose and follow-up on scoliosis treatment is to capture biplanar X-ray images and then use these to determine the sagittal frontal spinal curvature angles by the Cobb method. Reducing exposure to radiation is an important aspect for consideration, especially regarding children. The ZEBRIS spinal examination method is an external, non-invasive measurement method that uses an ultrasound-based motion analysis system. The aim of this study is to compare angle values of patients with adolescent idiopathic scoliosis (AIS) determined by the ZEBRIS spine examination method with the angle values defined by the gold standard Cobb method on biplanar X-ray images. Methods Subjects included 19 children with AIS (mean age 14.5±2.1 years, range 8–16 years, frontal plane thoracic Cobb angle 19.95±10.23°, thoracolumbar/lumbar angle 16.57±10.23°). The thoracic kyphosis and lumbar lordosis in the sagittal plane and the thoracic and lumbar scoliosis values were calculated by the Cobb method on biplanar X-ray images. The sagittal frontal spinal curvature angles were calculated from the position of the processus spinosus of 19 vertebrae, as determined by the ZEBRIS spine examination method. The validity of the ZEBRIS spine examination method was evaluated with Bland-Altman analyses between the sagittal and frontal spinal curvature parameters calculated from data determined by the ZEBRIS spine examination method and data obtained by the Cobb method on the X-ray images. Results and discussion Thoracic spinal curvature angles in sagittal and in frontal planes can be measured with sufficient accuracy. The slopes of the linear regression lines for thoracic kyphosis (TK) and thoracic scoliosis (TSC) are close to one (1.00 and 0.79 respectively), and the intercept values are below 5 degrees. The correlation between the TK and TSC values determined by the two methods is significant (p = 0.000) and excellent (rTK = 0.95, rTSC = 0.85). The differences are in the limit of agreement. The lumbar lordosis (LL) in the sagittal plane shows a very good correlation (rLL = 0.76); however the differences between the angles determined by the two methods are out of the limit of agreement in patients with major lumbar lordosis (LL≥50°). The thoracolumbar/lumbar spinal curvature angles in the frontal plane determined by ZEBRIS spine examination were underestimated at curvatures larger than 15°, mainly due to the rotational and pathological deformities of the scoliotic vertebrae. However, the correlation between lumbar scoliosis (LSC) values determined by the two methods is significant (p = 0.000) and excellent (rLSC = 0.84), the slopes are below one (0.71), the intercept values are below 5 degrees, and the differences between the angles determined by the two methods are within the limits of agreement. We could conclude that ZEBRIS spinal examination is a valid and reliable method for determination of sagittal and frontal curvatures during the treatment of patients with scoliosis. However, it cannot replace the biplanar X-ray examination for the visualization of spinal curvatures in the sagittal and frontal planes and the rotation of vertebral bodies during the diagnosis and annual evaluation of the progression.

Does bad posture affect the standing balance

Abstract Introduction: Bad posture is a well-known problem in children and adolescents, and it has a negative effect in adulthood. It can be hypothesized that due to bad posture, changes in the body’s position cause changes in standing balance. Objective: The objective of the study is to determine the influence of bad posture on the standing balance of school-aged children based on independent time–distance- and frequency-based foot centre-of-pressure parameters. Subjects and Methods: Subjects included 171 children (113 with neutral posture (70 boys and 73 girls), mean age: 10.7 ± 1.1 years (range: 9–13), and 68 with bad posture (22 boys and 46 girls), mean age: 10.7 ± 1.2 (range: 9–13)). The parameters were derived from the motion of the centre of pressure on a platform equipped with pressure sensors, on which the subjects were standing for 60 s with both feet and open eyes. Results: When comparing the two groups, the load distribution difference between the legs and the medium–high-frequency band power ratio in the mediolateral direction showed a significant difference out of 17 centre-of-pressure parameters. However, the other 15 parameters did not show any significant differences. Conclusion: There is no clearly significant degradation of postural control in children with bad posture, as the effects of altered posture are continuously corrected by the central nervous system. The asymmetric load between the two sides may further degrade muscular imbalance; thus, correcting bad posture is an important task of physiotherapy.

Importance of the Sequence-Directed DNA Shape for Specific Binding Site Recognition by the Estrogen-Related Receptor

Most nuclear receptors (NRs) bind DNA as dimers, either as hetero- or as homodimers on DNA sequences organized as two half-sites with specific orientation and spacing. The dimerization of NRs on their cognate response elements (REs) involves specific protein–DNA and protein–protein interactions. The estrogen-related receptor (ERR) belongs to the steroid hormone nuclear receptor (SHR) family and shares strong similarity in its DNA-binding domain (DBD) with that of the estrogen receptor (ER). In vitro, ERR binds with high affinity inverted repeat REs with a 3-bps spacing (IR3), but in vivo, it preferentially binds to single half-site REs extended at the 5′-end by 3 bp [estrogen-related response element (ERREs)], thus explaining why ERR was often inferred as a purely monomeric receptor. Since its C-terminal ligand-binding domain is known to homodimerize with a strong dimer interface, we investigated the binding behavior of the isolated DBDs to different REs using electrophoretic migration, multi-angle static laser light scattering (MALLS), non-denaturing mass spectrometry, and nuclear magnetic resonance. In contrast to ER DBD, ERR DBD binds as a monomer to EREs (IR3), such as the tff1 ERE-IR3, but we identified a DNA sequence composed of an extended half-site embedded within an IR3 element (embedded ERRE/IR3), where stable dimer binding is observed. Using a series of chimera and mutant DNA sequences of ERREs and IR3 REs, we have found the key determinants for the binding of ERR DBD as a dimer. Our results suggest that the sequence-directed DNA shape is more important than the exact nucleotide sequence for the binding of ERR DBD to DNA as a dimer. Our work underlines the importance of the shape-driven DNA readout mechanisms based on minor groove recognition and electrostatic potential. These conclusions may apply not only to ERR but also to other members of the SHR family, such as androgen or glucocorticoid, for which a strong well-conserved half-site is followed by a weaker one with degenerated sequence.

Erratum to: The Assessment of the Spinal Curvatures in the Sagittal Plane of Children Using an Ultrasound-Based Motion Analysing System[Annals of Biomedical Engineering, (2014), DOI: 10.1007/s10439-014-1160-z]

MÁRIA TAKÁCS, ERVIN RUDNER, ATTILA KOVÁCS, ZSANETT ORLOVITS, and RITA M. KISS MÁV Hospital Department of Orthopaedics, Verseghy Street 6-8, 5000 Szolnok, Hungary; MÁV Hospital Department of Rheumatology, Verseghy Street 6-8, 5000 Szolnok, Hungary; Institute of Mathematics, Budapest University of Technology and Economics, Egry József Street 1, 1111 Budapest, Hungary; and Cooperation Research Center for Biomechanics, Department of Mechatronics, Optics and Mechanical Engineering Informatics, Budapest University of Technology and Economics, M} uegyetem Rkp. 3-9, 1111 Budapest, Hungary