Exclusion of PINK1 as candidate gene for the late-onset form of Parkinson's disease in two European populations

BACKGROUND: Parkinson's disease (PD) is the second most common neurodegenerative disorder. Recently, mutations in the PINK1 (PARK6) gene were shown to rarely cause autosomal-recessively transmitted, early-onset parkinsonism. In order to evaluate whether PINK1 contributes to the risk of common late-onset PD we analysed PINK1 sequence variations. A German (85 patients) and a Norwegian cohort (90 patients) suffering from late-onset PD were screened for mutations and single nucleotide polymorphisms (SNPs) in the PINK1 gene. Both cohorts consist of well-characterized patients presenting a positive family history of PD in ~17%. Investigations were performed by single strand conformation polymorphism (SSCP), denaturating high performance liquid chromatography (DHPLC) and sequencing analyses. SNP frequencies were compared by the χ(2 )test RESULTS: Several common SNPs were identified in our cohorts, including a recently identified coding variant (Q115L) in exon 1. Genotyping of the Q115L variation did not reveal significant frequency differences between patients and controls. Pathogenic mutations in the PINK1 gene were not identified, neither in the German nor in the Norwegian cohort. CONCLUSION: Sequence variation in the PINK1 gene appears to play a marginal quantitative role in the pathogenesis of the late-onset form of PD, in German and Norwegian cohorts, if at all

Investigating Crack Initiation and Propagation of Concrete in Restrained Shrinkage Circular/Elliptical Ring Test

The restrained ring test, which is recommended by AASHTO and ASTM, has been used for assessing the potential of early-age cracking of concrete and other cement-based materials. Recently, a novel elliptical ring test method has been proposed to replace the circular ring test method for the purpose of shortening ring test duration and observing crack initiation and propagation more conveniently. In order to explore the mechanism of this novel test method, a numerical model is developed to analyze crack initiation and propagation process in restrained concrete rings, in which the effect of concrete shrinkage is simulated by a fictitious temperature drop applied on concrete causing the same strain as that induced by shrinkage. First, an elastic analysis is conducted to obtain the circumferential stress contour of a concrete ring subject to restrained shrinkage. Combined with the fictitious crack model, a fracture mechanics method is introduced to determine crack initiation and propagation, in which crack resistance caused by cohesive force acting on fracture process zone is considered. Finite element analysis is carried out to simulate the evolution of stress intensity factor in restrained concrete rings subject to circumferential drying. Cracking age and position of a series of circular/elliptical concrete rings are obtained from numerical analyses which agree reasonably well with experimental results. It is found that the sudden drop of steel strain observed in the restrained ring test represents the onset of unstable crack propagation rather than crack initiation. The results given by the AASHTO/ASTM restrained ring test actually reflects the response of a concrete ring as a structure to external stimulation, in this case restrained concrete shrinkage.The financial support from the National Natural Science Foundation of China under the grants of NSFC 51478083 & 51421064, Engineering and Physical Sciences Research Council under the grant of EP/I031952/1, and the National Basic Research Program of China (973 Program, Grant No. 2015CB057703) is gratefully acknowledged

Minimum Free Energy Path of Ligand-Induced Transition in Adenylate Kinase

Large-scale conformational changes in proteins involve barrier-crossing transitions on the complex free energy surfaces of high-dimensional space. Such rare events cannot be efficiently captured by conventional molecular dynamics simulations. Here we show that, by combining the on-the-fly string method and the multi-state Bennett acceptance ratio (MBAR) method, the free energy profile of a conformational transition pathway in Escherichia coli adenylate kinase can be characterized in a high-dimensional space. The minimum free energy paths of the conformational transitions in adenylate kinase were explored by the on-the-fly string method in 20-dimensional space spanned by the 20 largest-amplitude principal modes, and the free energy and various kinds of average physical quantities along the pathways were successfully evaluated by the MBAR method. The influence of ligand binding on the pathways was characterized in terms of rigid-body motions of the lid-shaped ATP-binding domain (LID) and the AMP-binding (AMPbd) domains. It was found that the LID domain was able to partially close without the ligand, while the closure of the AMPbd domain required the ligand binding. The transition state ensemble of the ligand bound form was identified as those structures characterized by highly specific binding of the ligand to the AMPbd domain, and was validated by unrestrained MD simulations. It was also found that complete closure of the LID domain required the dehydration of solvents around the P-loop. These findings suggest that the interplay of the two different types of domain motion is an essential feature in the conformational transition of the enzyme

Significance of the parkin and PINK1 gene in Jordanian families with incidences of young-onset and juvenile parkinsonism

<p>Abstract</p> <p>Background</p> <p>Parkinson's disease is a progressive neurodegenerative disorder, where most cases are sporadic with a late onset. In rare incidences familial forms of early-onset parkinsonism occur, and when recessively inherited, cases are often explained by mutations in either the <it>parkin </it>(PARK2) or <it>PINK1 </it>(PARK6) gene or on exceptional occasions the <it>DJ-1 </it>(PARK7) or <it>ATP13A2 </it>(PARK9) gene. Recessively inherited deletions/duplications and point mutations in the <it>parkin </it>gene are the most common cause of early-onset parkinsonism known so far, but in an increasing number of studies, genetic variations in the serine/threonine kinase domain of the <it>PINK1 </it>gene are found to explain early-onset parkinsonism.</p> <p>Methods</p> <p>In this study all families were from a population with a high incidence of consanguinity. We investigated 11 consanguineous families comprising 17 affected with recessively inherited young-onset parkinsonism for mutations both in the <it>parkin </it>and <it>PINK1 </it>gene. Exons and flanking regions were sequenced, and segregation patterns of genetic variation were assessed in members of the respective families. An exon dosage analysis was performed for all exons in both genes.</p> <p>Results</p> <p>In the <it>parkin </it>gene, a three generation family was identified with an exon 4 deletion segregating with disease. Both affected were homozygous for the deletion that segregated on a haplotype that spanned the gene in a haplotype segregation analysis that was performed using additional markers. Exon dosage analysis confirmed the recessive pattern of inheritance with heterozygous deletions segregating in healthy family members. In the <it>PINK1 </it>gene we identified two novel putative pathogenic substitutions, P416R and S419P, located in a conserved motif of the serine/threonine kinase domain. Both substitutions segregated with disease in agreement with a recessive pattern of inheritance within respective families and both were present as homozygous in two affected each. We also discuss common polymorphisms in the two genes found to be co-segregating within families.</p> <p>Conclusion</p> <p>Our results further extend on the involvement of <it>PINK1 </it>mutations in recessive early-onset parkinsonism with clinical features similar to carriers of <it>parkin </it>mutations.</p

Allosteric activation transitions in enzymes and biomolecular motors: insights from atomistic and coarse-grained simulations

The chemical step in enzymes is usually preceded by a kinetically distinct activation step that involves large-scale conformational transitions. In simple enzymes this step corresponds to the closure of the active site; in more complex enzymes, such as biomolecular motors, the activation step is more complex and may involve interactions with other biomolecules. These activation transitions are essential to the function of enzymes and perturbations in the scale and/or rate of these transitions are implicated in various serious human diseases; incorporating key flexibilities into engineered enzymes is also considered a major remaining challenge in rational enzyme design. Therefore it is important to understand the underlying mechanism of these transitions. This is a significant challenge to both experimental and computational studies because of the allosteric and multi-scale nature of such transitions. Using our recent studies of two enzyme systems, myosin and adenylate kinase (AK), we discuss how atomistic and coarse-grained simulations can be used to provide insights into the mechanism of activation transitions in realistic systems. Collectively, the results suggest that although many allosteric transitions can be viewed as domain displacements mediated by flexible hinges, there are additional complexities and various deviations. For example, although our studies do not find any evidence for cracking in AK, our results do underline the contribution of intra-domain properties (e.g., dihedral flexibility) to the rate of the transition. The study of mechanochemical coupling in myosin highlights that local changes important to chemistry require stabilization from more extensive structural changes; in this sense, more global structural transitions are needed to activate the chemistry in the active site. These discussions further emphasize the importance of better understanding factors that control the degree of co-operativity for allosteric transitions, again hinting at the intimate connection between protein stability and functional flexibility. Finally, a number of topics of considerable future interest are briefly discussed

Three-dimensional structural dynamics and fluctuations of DNA-nanogold conjugates by individual-particle electron tomography

DNA base pairing has been used for many years to direct the arrangement of inorganic nanocrystals into small groupings and arrays with tailored optical and electrical properties. The control of DNA-mediated assembly depends crucially on a better understanding of three-dimensional structure of DNA-nanocrystal-hybridized building blocks. Existing techniques do not allow for structural determination of these flexible and heterogeneous samples. Here we report cryo-electron microscopy and negative-staining electron tomography approaches to image, and three-dimensionally reconstruct a single DNA-nanogold conjugate, an 84-bp double-stranded DNA with two 5-nm nanogold particles for potential substrates in plasmon-coupling experiments. By individual-particle electron tomography reconstruction, we obtain 14 density maps at ∼2-nm resolution. Using these maps as constraints, we derive 14 conformations of dsDNA by molecular dynamics simulations. The conformational variation is consistent with that from liquid solution, suggesting that individual-particle electron tomography could be an expected approach to study DNA-assembling and flexible protein structure and dynamics