Comprehensive Structural and Thermodynamic Analysis of Prefibrillar WT α-Synuclein and Its G51D, E46K, and A53T Mutants by a Combination of Small-Angle X-ray Scattering and Variational Bayesian Weighting.

The in solution synchrotron small-angle X-ray scattering SAXS technique has been used to investigate an intrinsically disordered protein (IDP) related to Parkinson's disease, the α-synuclein (α-syn), in prefibrillar diluted conditions. SAXS experiments have been performed as a function of temperature and concentration on the wild type (WT) and on the three pathogenic mutants G51D, E46K, and A53T. To identify the conformers that populate WT α-syn and the pathogenic mutants in prefibrillar conditions, scattering data have been analyzed by a new variational bayesian weighting method (VBWSAS) based on an ensemble of conformers, which includes unfolded monomers, trimers, and tetramers, both in helical-rich and strand-rich forms.

Superoxide Dismutases SOD1 and SOD2 Rescue the Toxic Effect of Dopamine-Derived Products in Human SH-SY5Y Neuroblastoma Cells.

The preferential loss of dopaminergic neurons in the substantia nigra pars compacta is one of the pathological hallmarks characterizing Parkinson's disease. Although the pathogenesis of this disorder is not fully understood, oxidative stress plays a central role in the onset and/or progression of Parkinson's disease and dopamine itself has been suggested to participate in the preferential neuronal degeneration through the induction of oxidative conditions. In fact, the accumulation of dopamine into the cytosol can lead to the formation of reactive oxygen species as well as highly reactive dopamine-quinones.

Superoxide dismutating molecules rescue the toxic effects of PINK1 and parkin loss.

Reactive oxygen species exert important functions in regulating several cellular signalling pathways. However, an excessive accumulation of reactive oxygen species can perturb the redox homeostasis leading to oxidative stress, a condition which has been associated to many neurodegenerative disorders. Accordingly, alterations in the redox state of cells and mitochondrial homeostasis are established hallmarks in both familial and sporadic Parkinson's disease cases.

High-Pressure-Driven Reversible Dissociation of α-Synuclein Fibrils Reveals Structural Hierarchy.

The analysis of the α-synuclein (aS) aggregation process, which is involved in Parkinson's disease etiopathogenesis, and of the structural feature of the resulting amyloid fibrils may shed light on the relationship between the structure of aS aggregates and their toxicity. This may be considered a paradigm of the ground work needed to tackle the molecular basis of all the protein-aggregation-related diseases. With this aim, we used chemical and physical dissociation methods to explore the structural organization of wild-type aS fibrils.

Recent findings on the physiological function of DJ-1: Beyond Parkinson's disease.

Several mutations in the gene coding for DJ-1 have been associated with early onset forms of parkinsonism. In spite of the massive effort spent by the scientific community in understanding the physiological role of DJ-1, a consensus on what DJ-1 actually does within the cells has not been reached, with several diverse functions proposed. At present, the most accepted function for DJ-1 is a neuronal protective role against oxidative stress.

Pressure effects on α-synuclein amyloid fibrils: An experimental investigation on their dissociation and reversible nature.

α-synuclein amyloid fibrils are found in surviving neurons of Parkinson's disease affected patients, but the role they play in the disease development is still under debate. A growing number of evidences points to soluble oligomers as the major cytotoxic species, while insoluble fibrillar aggregates could even play a protection role. In this work, we investigate α-synuclein fibrils dissociation induced at high pressure by means of Small Angle X-ray Scattering and Fourier Transform Infrared Spectroscopy.

Superoxide Dismutase (SOD)-mimetic M40403 Is Protective in Cell and Fly Models of Paraquat Toxicity: IMPLICATIONS FOR PARKINSON DISEASE.

Parkinson disease is a debilitating and incurable neurodegenerative disorder affecting ∼1-2% of people over 65 years of age. Oxidative damage is considered to play a central role in the progression of Parkinson disease and strong evidence links chronic exposure to the pesticide paraquat with the incidence of the disease, most probably through the generation of oxidative damage. In this work, we demonstrated in human SH-SY5Y neuroblastoma cells the beneficial role of superoxide dismutase (SOD) enzymes against paraquat-induced toxicity, as well as the therapeutic potential of the SOD-mimetic compound M40403.

LRRK2 phosphorylates pre-synaptic N-ethylmaleimide sensitive fusion (NSF) protein enhancing its ATPase activity and SNARE complex disassembling rate.

Background: Lrrk2, a gene linked to Parkinson's disease, encodes a large scaffolding protein with kinase and GTPase activities implicated in vesicle and cytoskeletal-related processes. At the presynaptic site, LRRK2 associates with synaptic vesicles through interaction with a panel of presynaptic proteins.

Results: Here, we show that LRRK2 kinase activity influences the dynamics of synaptic vesicle fusion.

Anti-Oxidants in Parkinson's Disease Therapy: A Critical Point of View.

Parkinson's disease (PD) is a degenerative neurological syndrome, which is characterized by the preferential death of dopaminergic (DAergic) neurons in the Substantia Nigra. The pathogenesis of this disorder remains poorly understood and PD is still incurable. Current drug treatments are aimed primarily for the treatment of symptoms to improve the quality of life.

Analysis of the Catecholaminergic Phenotype in Human SH-SY5Y and BE(2)-M17 Neuroblastoma Cell Lines upon Differentiation.

Human cell lines are often used to investigate cellular pathways relevant for physiological or pathological processes or to evaluate cell toxicity or protection induced by different compounds, including potential drugs. In this study, we analyzed and compared the differentiating activities of three agents (retinoic acid, staurosporine and 12-O-tetradecanoylphorbol-13-acetate) on the human neuroblastoma SH-SY5Y and BE(2)-M17 cell lines; the first cell line is largely used in the field of neuroscience, while the second is still poorly characterized. After evaluating their effects in terms of cell proliferation and morphology, we investigated their catecholaminergic properties by assessing the expression profiles of the major genes involved in catecholamine synthesis and storage and the cellular concentrations of the neurotransmitters dopamine and noradrenaline.

Synergistic and antagonistic effects of thermal shock, air exposure, and fishing capture on the physiological stress of Squilla mantis (Stomatopoda).

This study is aimed at assessing the effects of multiple stressors (thermal shock, fishing capture, and exposure to air) on the benthic stomatopod Squilla mantis, a burrowing crustacean quite widespread in the Mediterranean Sea. Laboratory analyses were carried out to explore the physiological impairment onset over time, based on emersion and thermal shocks, on farmed individuals. Parallel field-based studies were carried out to also investigate the role of fishing (i.

Are dopamine derivatives implicated in the pathogenesis of Parkinson's disease?

Parkinson's disease (PD) is the most common motor system disorder affecting 1-2% of people over the age of sixty-five. Although PD is generally a sporadic neurological disorder, the discovery of monogenic, hereditable forms of the disease, representing 5-10% of all cases, has been very important in helping to partially delineate the molecular pathways that lead to this pathology. These mechanisms include impairment of the intracellular protein-degradation pathways, protein aggregation, mitochondria dysfunction, oxidative stress and neuroinflammation.

Dysfunction of dopamine homeostasis: clues in the hunt for novel Parkinson's disease therapies.

Parkinson's disease is the second most common neurodegenerative disorder and, at present, has no cure. Both environmental and genetic factors have been implicated in the etiology of the disease; however, the pathogenic pathways leading to neuronal degeneration are still unclear. Parkinson's disease is characterized by the preferential death of a subset of neurons in the mesencephalon that use dopamine as neurotransmitter for synaptic communication.

Quaternary structure heterogeneity of oligomeric proteins: a SAXS and SANS study of the dissociation products of Octopus vulgaris hemocyanin.

Octopus vulgaris hemocyanin shows a particular self-assembling pattern, characterized by a hierarchical organization of monomers. The highest molecular weight aggregate is a decamer, the stability of which in solution depends on several parameters. Different pH values, buffer compositions, H₂O/D₂O ratios and Hofmeister's salts result in modifications of the aggregation state of Octopus vulgaris hemocyanin.

QUAFIT: a novel method for the quaternary structure determination from small-angle scattering data.

The new QUAFIT method for determining the quaternary structure of biological macromolecular assemblies by analyzing x-ray or neutron small-angle scattering data is presented. The method is based on the idea that asymmetric monomers, formed by rigid domains of known atomic structure possibly connected by flexible linkers of known sequence, are assembled according to a point-group symmetry combined with a screw axis. Scattering amplitudes of domains and linkers are determined by means of a spherical harmonics expansion and combined to get the form factor of the assembly.

Human SOD2 modification by dopamine quinones affects enzymatic activity by promoting its aggregation: possible implications for Parkinson's disease.

Mitochondrial dysfunction and oxidative stress are considered central in dopaminergic neurodegeneration in Parkinson's disease (PD). Oxidative stress occurs when the endogenous antioxidant systems are overcome by the generation of reactive oxygen species (ROS). A plausible source of oxidative stress, which could account for the selective degeneration of dopaminergic neurons, is the redox chemistry of dopamine (DA) and leads to the formation of ROS and reactive dopamine-quinones (DAQs).

Insights into the oligomerization process of the C-terminal domain of human plasma membrane Ca²+-ATPase.

Plasma membrane calcium pumps (PMCAs) sustain a primary transport system for the specific removal of cytosolic calcium ions from eukaryotic cells. PMCAs are characterized by the presence of a C-terminal domain referred to as a regulatory domain. This domain is target of several regulatory mechanisms: activation by Ca²+-calmodulin complex and acidic phospholipids, phosphorylation by kinase A and C, proteolysis by calpain and oligomerization.

Structural basis of the lactate-dependent allosteric regulation of oxygen binding in arthropod hemocyanin.

Hemocyanin (Hc) is an oxygen carrier protein in which oxygen binding is regulated by allosteric effectors such as H(+) and L-lactate. Isothermal titration calorimetric measurements showed that L-lactate binds to dodecameric and heterohexameric Hc and to the CaeSS3 homohexamer but not to the CaeSS2 monomer. The binding of lactate caused no change in the optical absorption and x-ray absorption spectra of either oxy- or deoxy-Hc, suggesting that no structural rearrangement of the active site occurred.

Solution structures of 2×6-meric and 4×6-meric hemocyanins of crustaceans Carcinus aestuarii, Squilla mantis and Upogebia pusilla.

Arthropod hemocyanins (Hcs) are a family of large, high molecular mass, extracellular oxygen transport proteins. They form oligomeric quaternary structures based on different arrangements of a basic 6×75 kDa hexameric unit. Their complex quaternary structures present binding sites for allosteric effectors and regulate the oxygen binding process in a cooperative manner.

Cytosolic carbonic anhydrase in the Gulf toadfish is important for tolerance to hypersalinity.

Carbonic anhydrase (CA) is a ubiquitous enzyme involved in acid-base regulation and osmoregulation. Many studies have demonstrated a role for this enzyme in fish osmoregulation in seawater as well as freshwater. However, to date CA responses of marine fish exposed to salinities exceeding seawater (approximately 35 ppt) have not been examined.

Unusual oxygen binding behavior of a 24-meric crustacean hemocyanin.

Hemocyanins from Crustacea usually are found as 1x6 or 2x6-meric assemblies. An exception is the hemocyanin isolated from thalassinidean shrimps where the main component is a 24-meric structure. Our analysis of oxygen binding data of the thalassinidean shrimp Upogebia pusilla based on a three-state MWC-model revealed that despite the 24-meric structure the functional properties can be described very well based on the hexamer as allosteric unit.

Seasonal variation of trehalose and glycerol concentrations in winter snow-active insects.

Different kinds of molecules were identified as antifreezing agents in the body fluids of cold tolerant invertebrates: sugars, polyols and proteins. While none of the active arthropods were so far reported to accumulate polyols, these compounds are present in the haemolymph of species that hibernate in a passive stage such as diapause. In this work we investigated insect species that are active during winter and we demonstrated the ability of the mecopteran Boreus hiemalis (Mecoptera, Boreidae), the wingless fly Chionea sp.

Molecular basis of the Bohr effect in arthropod hemocyanin.

Flash photolysis and K-edge x-ray absorption spectroscopy (XAS) were used to investigate the functional and structural effects of pH on the oxygen affinity of three homologous arthropod hemocyanins (Hcs). Flash photolysis measurements showed that the well-characterized pH dependence of oxygen affinity (Bohr effect) is attributable to changes in the oxygen binding rate constant, k(on), rather than changes in k(off). In parallel, coordination geometry of copper in Hc was evaluated as a function of pH by XAS.

Respiration rate and oxy-regulatory capacity in cold stenothermal chironomids.

The effects of temperature and oxygen saturation on the respiration rate of two cold stenothermal chironomids, Diamesa insignipes and Pseudodiamesa branickii were investigated. Fourth instar larvae were collected in winter in a glacio-rhithral stream (1300 m a.s.

Role of the tertiary structure in the diphenol oxidase activity of Octopus vulgaris hemocyanin.

The functional differences between the oxygen transport protein Hemocyanin and the enzymes Tyrosinase and Catechol oxidase are believed to be governed, at least in part, by the tertiary structure, which differs in these molecules and controls the accessibility of their copper containing active site for substrate(s). Accordingly, Octopus vulgaris Hemocyanin catalyses the o-diphenol oxidation to o-quinone at a very low rate. The crystallographic structure of one of the functional units (called Odg) of O.